Dosage regimen for administrating a CD19×CD3 bispecific antibody to patients at risk for potential adverse effects

ABSTRACT

A method for assessing the risk of potential adverse effects for a human patient receiving a CD19×CD3 bispecific antibody is provided. The method comprises determining the total B count in the patient, and identifying a B cell number indicative of a patient at risk of potential adverse effects from the antibody. The method further provides a dosing schedule for administering the antibody to the patient identified as at risk of potential adverse effects. Also provided is a pharmaceutical package or kit comprising a first dose and a second dose, and optionally a third dose, of the CD19×CD3 bispecific antibody as defined in the methods/dosage regimen of the disclosure.

The present invention relates to a method for assessing (analyzing) therisk of potential adverse effects for a human patient mediated by theadministration of a CD19×CD3 bispecific antibody to said patientcomprising determining the ratio of B cells to T cells or total B cellsof said patient, wherein a ratio of about 1:5 or lower and/or a total Bcell count of less than about 50 B cells per microliter of peripheralblood is indicative for a risk of potential adverse effects for saidpatient. Accordingly, the present invention relates to a method (dosageregimen) for administering a CD19×CD3 bispecific antibody to a humanpatient having a B:T cell ratio of about 1:5 or lower or a total B cellcount of less than about 50 B cells per microliter in peripheral blood,comprising (a) administering a first dose of said antibody for a firstperiod of time; and consecutively (b) administering a second dose ofsaid antibody for a second period of time, wherein said second doseexceeds said first dose. In some embodiments, a third dose of saidantibody is optionally administered for a third period of time or forthe same time period as the first and/or second doses. The third dose ofsaid antibody exceeds said second dose. This dosage regimen can beapplied in methods for treating malignant CD19 positive lymphocytes orfor ameliorating and/or preventing an adverse effect mediated by theadministration of said bispecific antibody. The present invention alsorelates to the use of a CD19×CD3 bispecific antibody for the preparationof a pharmaceutical composition to be used in a method of the presentinvention. A pharmaceutical package or kit comprising a first dose and asecond dose and optionally a third dose of said antibody as defined inthe methods/dosage regimen of the present invention is disclosed aswell.

Antibody-based cancer therapies require a target antigen firmly bound tothe surface of cancer cells in order to be active. By binding to thesurface target, the antibody can directly deliver a deadly signal to thecancer cell or indirectly by, for example, recruiting a cytotoxic Tcell, if it is a bispecific antibody. In an ideal treatment scenario, atarget antigen is abundantly present and accessible on every cancer celland is absent, shielded or much less abundant on normal cells. Thissituation provides the basis for a therapeutic window in which a definedamount of the antibody-based therapeutic effectively hits cancer cellsbut spares normal cells.

Though antibodies are an effective means in treating many disorders, inparticular cancer, their administration is not necessarily devoid ofside effects. Adverse effects may cause a reversible or irreversiblechange in the health status of a patient. As adverse effects could beharmful and undesired, it is highly desirable to avoid them. However,though it is known that a medicament can cause adverse effects, itsprescription and administration could not be avoided or is accepted,since the medicament has an outstanding beneficial therapeutic effect ormay even be life-saving.

In clinical trials, a general distinction can be made between adverseeffects (AEs) and serious adverse effects (SAEs). Specifically, adverseeffects can be classified in 5 grades in accordance with the CommonTerminology Criteria for Adverse Events (CTCAE). Grade 1 relates to mildAE, Grade 2 to moderate AE, Grade 3 to severe AE, Grade 4 tolife-threatening or disabling AE, while Grade 5 means death related toAE.

An adverse effect observed in antibody therapy is the occurrence ofinfusion-related side effects, such as the cytokine release syndrome(“CRS”). Other adverse side effects described to be associated with CRSare fatigue, vomiting, tachycardia, hypertension, back pain, but alsocentral nervous system reactions (CNS reactions), such as seizures,encephalopathy, cerebral edema, aseptic meningitis, and headache.

Cytokine release and neurological reactions have not only been observedwith monoclonal antibodies binding to the T cell receptor but also witha CD19×CD3 bispecific single chain antibody binding to the CD3 part ofthe T cell receptor (called Blinatumomab (MT103)).

Blinatumomab (MT103) is a lymphoma-directed, recombinant bispecificsingle-chain CD19×CD3 antibody that binds to CD19 on the surface ofalmost all B cells and B tumor cells and concomitantly can engage a Tcell, thereby triggering the T-cell to kill the target B cell or B tumorcell. Blinatumomab consists of four immunoglobulin variable domainsassembled into a single polypeptide chain. Two of the variable domainsform the binding site for CD19, a cell surface antigen expressed on mostB cells and B tumor cells. The other two variable domains form thebinding site for the CD3 complex on T cells. Blinatumomab is designed todirect the body's cytotoxic, or cell-destroying, T cells against tumorcells, and represent a new therapeutic approach to cancer therapy.Blinatumomab is presently in clinical trials.

As described for instance in WO 99/54440, adverse effects have beenobserved in a previous study performed with Blinatumomab applied inrepeated bolus infusions to a patient with B-cell derived chroniclymphatic leukaemia (B-CLL). As shown in FIGS. 19 and 20 of WO 99/54440,release of TNF, IL-6 and IL-8 has been found in response to each of thetwo administered 20 minute-infusions of 3 microgram and 10 microgram ofthe mentioned bispecific single chain antibody, respectively, withcytokine release after each administration. Maximal cytokine release wasobserved after administration of 10 microgram of bispecific single chainantibody. In a following clinical trial study, in which escalating dosesof the CD19×CD3 bispecific single chain antibody have been administeredto patients with B cell malignancies as bolus infusions, adverse effectshave also been observed. According to a retrospective analysis, 7 out of22 patients showed an early neurological reaction, including, forexample, confusion, ataxia, speech disorder, or disorientation.

In order to try to better manage these undesired side effects, the modeof administration of the CD19×CD3 bispecific single chain antibody hasbeen changed in that it has been switched over from bolus infusion to acontinuous intravenous administration of said antibody for a longerperiod of time. As shown in Bargou et al. (Science 321 (2008): 974-7),doses as low as 0.005 milligrams per square meter per day continuouslyadministered to non-Hodgkin's lymphoma patients over four weeks led toan elimination of lymphoma target cells in blood. Partial and completetumor regressions were first observed at a dose level of 0.015milligrams/m²/d, and all seven patients treated at a dose level of 0.06milligrams/m²/d experienced a tumor regression (Bargou et al., citedabove). The CD19×CD3 bispecific single chain antibody also led toclearance of tumor cells from bone marrow and liver. However, thoughthis (still ongoing) study established clinical proof of concept for thetherapeutic potency of the CD19×CD3 bispecific single chain antibodyformat in the treatment of blood-cell derived cancer, neurologicalreactions have been found in the course of the aforementioned clinicaltrial. Accordingly, since Blinatumomab is a very promising candidatemedicament for treating non-Hodgkin's lymphoma (NHL), acutelymphoblastic leukemia (ALL), chronic lymphocytic leukemia (CLL) and/ormantle cell lymphoma, it is highly desirable to reduce or evencompletely avoid undesired side-effects in the treatment of patients inneed thereof with the CD19×CD3 bispecific single chain antibody.

Evidently, it is difficult to design a CD19×CD3 antibody-based therapy,which does not cause CNS (neurological) reactions including neurologicalreactions, or, to put it differently, it is desired to provide aCD19xCD13 antibody-based medical therapies with increased patienttolerability, i.e., reduced or even no undesired adverse effects such asCNS reactions.

Though pharmaceutical means and methods which allow a more gradualactivation of T cell populations (see WO 2007/068354) already helped toavoid significant adverse side effects in patients treated with theCD19×CD3 bispecific single chain antibody, neurological reactions couldunfortunately not be prevented by these measures, in particular in casesin which doses of more than 5 to 10 microgram per square meter per day(i.e. 24 h) of the antibody have been administered.

Thus, the technical problem underlying the present invention was toprovide methods for assessing the risk of potential adverse effects fora human patient mediated by the administration of a CD19×CD3 bispecificantibody to said patient in order to then devise dosage regimens andmethods to overcome the above problem.

The present invention addresses this need and thus provides embodimentsconcerning methods as well as dosage regimens for administering aCD19×CD3 bispecific antibody to a human patient.

These embodiments are characterized and described herein and reflectedin the claims.

It must be noted that as used herein, the singular forms “a”, “an”, and“the”, include plural references unless the context clearly indicatesotherwise. Thus, for example, reference to “a reagent” includes one ormore of such different reagents and reference to “the method” includesreference to equivalent steps and methods known to those of ordinaryskill in the art that could be modified or substituted for the methodsdescribed herein.

All publications and patents cited in this disclosure are incorporatedby reference in their entirety. To the extent the material incorporatedby reference contradicts or is inconsistent with this specification, thespecification will supersede any such material.

Unless otherwise indicated, the term “at least” preceding a series ofelements is to be understood to refer to every element in the series.Those skilled in the art will recognize, or be able to ascertain usingno more than routine experimentation, many equivalents to the specificembodiments of the invention described herein. Such equivalents areintended to be encompassed by the present invention.

Throughout this specification and the claims which follow, unless thecontext requires otherwise, the word “comprise”, and variations such as“comprises” and “comprising”, will be understood to imply the inclusionof a stated integer or step or group of integers or steps but not theexclusion of any other integer or step or group of integer or step.

Several documents are cited throughout the text of this specification.Each of the documents cited herein (including all patents, patentapplications, scientific publications, manufacturer's specifications,instructions, etc.), whether supra or infra, are hereby incorporated byreference in their entirety. Nothing herein is to be construed as anadmission that the invention is not entitled to antedate such disclosureby virtue of prior invention.

In view of the adverse events, particularly the CNS events includingneurological reactions observed with antibodies, also including theCD19×CD3 bispecific antibody, the finding that the CD19×CD3 bispecificsingle chain antibody can be administered so that it is tolerated by thepatients, if it is administered in accordance with the dosage regimen asprovided herein, is definitely remarkable.

Specifically, the present inventors observed that those patients, towhom a CD19×CD3 bispecific antibody was administered, encountered CNSevents, if they had a B:T cell ratio of about 1:5 or lower or a total Bcell count of less than about 50 B cells per microliter of peripheralblood. Accordingly, the present invention for the first time establishesa low B:T cell ratio or a low total B cell count as a potential highrisk factor for the occurrence of adverse effects including neurologicalreactions in the treatment of malignant CD19 positive lymphocytesoccurring in leukemias and lymphomas (see Examples 2, 3 and 4).

Particularly, the inventors of the present application observed thatnon-Hodgkin lymphoma (NHL) patients and acute lymphoblastic leukemia(ALL) patients with a low B:T cell ratio or a low total B cell count inperipheral blood have an increased risk for the development of an earlyneurological reaction. This neurological reaction occurs mainly duringthe first day(s) of treatment with a CD19xCD13 bispecific antibody. Inparticular, the majority of the neurological reactions occurred afterabout 12 to 120 hours after start of treatment. These neurologicalreactions were transient, fully reversible and resolved without sequelaewithin 3 to 72 hours after stop of the treatment. The inventors madethese unexpected observations in various clinical trial studies usingthe CD19×CD3 bispecific antibody:

Looking at “short-term” (bolus) infusion trials, 7 out of 22 patientshad an early neurological reaction. 6 of these 7 patients had a low B:Tcell ratio, i.e., a B:T cell ratio of about 1:5 or lower, beforetreatment. Of the remaining 15 patients without neurological reaction,only 1 patient had a low B:T cell ratio.

In an NHL clinical trial (see Bargou et al., cited above), a total of 39patients have been treated until August 2008. At this time point, it hasbeen found that all patients with a neurological reaction that led topermanent discontinuation of the CD19×CD3 bispecific antibody treatmenthad a low B:T cell ratio (i.e., a B:T cell ratio threshold below 1:5).In particular, 5 neurological reactions have been observed in 10patients with low B:T cell ratio (5/10), while no patient with a highB:T cell ratio (i.e., a B:T cell ratio higher than 1:5) had aneurological reaction that would have led to permanent discontinuationof CD19×CD3 bispecific antibody treatment (0/29).

Thereafter, a specific cohort for patients with low B:T cell ratioand/or a lower total B cell count i.e. an increased risk for earlyneurological reactions, was established in order to prospectivelyanalyze the outlined theory and to specifically find mitigation stepsfor the patients at increased risk.

Since establishing these separated cohorts for high risk patients, 8 NHLpatients were prospectively treated (data as of July 2009): 6 patientswith low B:T cell ratio, 2 patients with high B:T cell ratio. Again nopatient with a high B:T cell ratio had a neurological reaction, while 3out of 6 patients with a low B:T cell ratio had a neurological reaction,leading to discontinuation of the treatment. In sum, 69 NHL patients,including B-cell chronic lymphocytic leukemia (CLL) and mantle celllymphoma (MCL), have been treated with a CD19×CD3 bispecific antibody,both with bolus infusion and continuous infusion:

Neurological reactions have been observed in 61% of the patients withlow B:T cell ratio. In contrast, only 2% of the patients with high B:Tcell ratio showed such adverse events (see the following examples).

In another clinical trial phase II study, 15 μg of CD19×CD3 bispecificsingle chain antibody per square meter patient body surface area per dayhave been administered to adult ALL patients by continuous infusion forat least four weeks. One out of 11 ALL patients of the high risk grouphaving a B:T cell ratio below 1:5 showed a neurological reaction,leading to discontinuation of the treatment. In contrast, none of the 6patients of the low risk group having a B:T cell ratio higher than 1:5showed a neurological reaction.

Moreover, in a retrospective analysis of 39 NHL patients, a baseline Bcell to T cell (B:T) ratio in peripheral blood at or below 1:5 to 1:10was identified as the only predictive factor for the subsequentoccurrence of neurological AEs. The predictive value was thenprospectively confirmed in 8 additional patients (see Example 1).

These data establish a low B:T cell ratio, i.e., a B:T cell ratio ofabout 1:5 or lower as a potential high risk factor for the occurrence ofadverse effects including neurological reactions in the treatment ofmalignant CD19 positive lymphocytes occurring in leukemia and lymphomasuch as NHL, MCL, CLL and ALL in patients who are treated with aCD19×CD3 bispecific antibody (see Examples 1 and 4).

In addition to utilizing B:T cell ratios as a predictor of CNS events ina patient, it has been determined that initial total B cell counts inthe peripheral blood is also a good predictor of CNS events in apatient. In continuing clinical trials, a total of 89 patientsundergoing treatment with blinatumomab (70 patients with NHL and 19patients with ALL) were evaluated. Of these 89 patients, 19 patients hada permanent discontinuation from treatment with blinatumomab due to aCNS event; and low B cell counts of less than about 50 B cells permicroliter of peripheral blood in individual patients was useful inidentifying these patients susceptible to a CNS event, and correlatedwith low B:T cell ratios.

More specifically, when all evaluable patients are analyzed by themedian of their individual initial B cell count in peripheral blood(median 42.5/microliter), 16 out of 44 patients below this median had aCNS event (36.4%), while 3 out of 44 patients above this median had aCNS event (6.8%); of note: the one patient that represents the medianhad no CNS event.

Further analyses of this data provide usable cut-offs for separatingpatients at risk for CNS events leading to permanent discontinuationfrom blinatumomab treatment based on the initial total B cell count. Inview of this data, it has been determined that a total B cell count ofless than 35/microliter identifies 16 out of 40 patients with a low Bcell count (<35/μL) that showed a CNS event (35.6%) while 3 out of 49patients with a high B cell count (>35/μl) showed a CNS event (6.1%).

Of note, all 3 patients with the CNS events in the group with high totalB cell counts had specifics that distinguish them from the majority ofother patients. For example, 2 patients had received the highest dose of90 μg/m²/d which was determined as exceeding the MTD as a starting doseand the third patient discontinued treatment only after a break and are-start of blinatumomab (this was not considered a new treatment cycleand therefore the initial B cell count was used and not the B cell countbefore the re-start). If the total B cell count before re-start wouldhave been used this patient would also have fallen into the group ofpatients with a B cell count below 35/μl.

Similarly considering a cut-off to a B cell count of about 50/microliter(50/μl) to assess the risk of adverse effects is almost identical withthe median as cut-off; i.e.: 16 out of 45 patients with a low B cellcount (<50/μL) showed a CNS event (40.0%) while 3 out of 44 patientswith a high B cell count (>50/μl) showed a CNS event (6.7%). See FIG. 1for a Summary of this information.

Similarly, the median total B cell counts before blinatumomab treatmentstarted were 113.25/μl for all patients without CNS event leading topermanent discontinuation and 1.12/μl for all patients with such a CNSevent.

Therefore, a low peripheral B cell count before treatment start is agood predictor for the occurrence of CNS events leading to permanentdiscontinuation of blinatumomab treatment, and vice versa, a highperipheral total B cell count before treatment started is a goodpredictor for a reduced occurrence of CNS events leading to permanentdiscontinuation of blinatumomab treatment (see FIGS. 2A and 2B showingdata from two clinical studies applying a CD19×CD3 bispecific antibody).Low total B cell counts of less than 40-50/microliter correlate wellwith the low B:T cell ratio of less than about 1:8-1:9 and indicates apatient at risk for the occurrence of a CNS event during treatment withblinatumomab.

Thus, it was an aim of the present invention to provide a method thatallows identifying patients who may be at a risk of suffering fromadverse effects when being treated with a CD19×CD3 bispecific antibody.This method will improve drug compliance, since the identification ofpatients who are at a risk of suffering from adverse effects allowsadjusting the dosage regimen of the CD19×CD3 bispecific antibody. Infact, the present inventors have applied their finding that a B:T cellratio of about 1:5 or lower and/or a total B cell count of less thanabout 50 B cells/microliter of peripheral blood could be a potentialrisk factor for suffering from adverse effects in the treatment with aCD19×CD3 bispecific antibody and have thus developed a dosage regimenwhich is intended to prevent and/or ameliorate these adverse effects.

Accordingly, in a first aspect the present invention provides a methodfor assessing (analysing) the risk of potential adverse effects for ahuman patient mediated by the administration of a CD19×CD3 bispecificantibody to said patient comprising determining in a sample from saidpatient the ratio of B cells to T cells of said patient, wherein a ratioof about 1:5 or lower and/or a total B cell count of less than about 50B cells/microliter of peripheral blood is indicative for a risk ofpotential adverse effects for said patient. On the other hand, a ratioof higher than 1:5 and/or a total B cell count of greater than about 50B cells/microliter of peripheral blood is indicative for a decreasedrisk of potential adverse effects for said patients. Thus, the methodsof the present invention allow a “grouping” of patients into low andhigh risk patients. Dependent on the risk group, a patient-tailoredtreatment regimen should be applied, as described herein.

“Assessing (analysing) the risk” means that the method of the firstaspect of the present invention aims at assessing or analysing as towhether or not a patient has a higher or lower likelihood or probability(i.e., an increased or decreased risk, respectively) to encounteradverse effects. Accordingly, as is commonly known, a risk does notnecessarily mean that a patient will or will not encounter adverseeffects.

In the present invention, when a patient has a B:T cell ratio of about1:5 or lower and/or a total B cell count of less than about 50 Bcells/microliter of peripheral blood, said patient has (is at) anincreased risk of potential adverse effects, also including the onset ofan adverse effect, while a patient who has a B:T cell ratio higher than1:5 and/or a total B cell count of greater than about 50 Bcells/microliter of peripheral blood does not have (is not at) or atleast has (is at) a decreased risk of potential adverse effects, alsoincluding the onset of an adverse effect.

Accordingly, a B:T cell ratio of about 1:5 or lower is indicative for arisk of adverse effects, while a B:T cell ratio of higher than 1:5 isnot indicative for a risk of adverse effects. Similarly, a total B cellcount of less than about 50 B cells/microliter of peripheral blood isindicative for a risk of adverse effects, while a total B cell count ofgreater than 50 B cells/microliter of peripheral blood is not indicativefor a risk of adverse effects.

Thus, the term “indicative for” when used in the context of the methodof the first aspect of the present invention means that a patient has anincreased risk of potential adverse effects if the B:T cell ratio isabout 1:5 or lower and/or if the total B cell count is less than about50 B cells/microliter of peripheral blood or has a decreased risk ofpotential adverse effects if the B:T cell ratio is higher than 1:5and/or a total B cell count of greater than 50 B cells/microliter ofperipheral blood.

An “adverse effect” is a harmful and undesired effect resulting frommedication in the treatment of a patient with a CD19×CD3 bispecificantibody. An adverse effect may also be termed a “side effect”. Someadverse effects only occur only when starting, increasing ordiscontinuing a treatment. The inventors have observed that the adverseeffect seen in the treatment of patients with a CD19×CD3 bispecificantibody occurred after about 12 to 120 hours after the start of thetreatment and are reversible.

An adverse effect may cause medical complications. The inventors haveobserved neurological reactions in patients treated with a CD19×CD3bispecific antibody. These neurological reaction, unless they can bestopped or avoided, lead to non-compliance with the CD19×CD3 bispecificantibody treatment.

However, as mentioned herein, the inventors found that the B:T cellratio and/or a total B cell count is an indicator as to whether or notpatients are at a risk of potential adverse side effects. Specifically,a B:T cell ratio of about or lower than 1:5 and/or a total B cell countof less than about 50 B cells/microliter of peripheral blood is anindicator that patients are at a risk of potential side effects, while aB:T cell ratio higher than about 1:5 and/or a total B cell count ofgreater than about 50 B cells/microliter of peripheral blood is anindicator that patients have no or at least have a decreased risk ofpotential side.

As mentioned before, the method of the first aspect of the presentinvention is for assessing (analysing) the risk of adverse effects and arisk includes the assessment/analysis of likelihood or a probability.Accordingly, the term “potential” when used in the context of adverseeffects means that—though a patient may have a B:T cell ratio of about1:5 or lower and/or a total B cell count of less than about 50 Bcells/microliter of peripheral blood—said patient does not necessarilyhave to encounter adverse effects.

Likewise, though a patient may have a B:T cell ratio higher than about1:5 or a total B cell count of greater than about 50 B cells/microliterof peripheral blood—said patient does not necessarily have to notencounter adverse effects. Accordingly, the term “potential” impliesthat the method of the first aspect of the present invention providespredictions as to whether or not a patient may encounter adverseeffects, but—self-explanatory as it is—cannot provide a 100% safeprediction, since, apart from the B:T cell ratio or total B cell count,individual factors such as sex, age, weight, nutritional status, healthstatus, pre-medication etc. may have an influence as to whether or not apatient will encounter adverse effects.

In accordance with the present invention an adverse effect is preferablycharacterized by a neurological reaction (also sometimes referred toherein as “CNS reaction” or “CNS event”, for which reason these termscan be equally used). Said neurological reaction is preferably one ormore selected from the group consisting of: confusion, ataxia,disorientation, dysphasia, aphasia, speech impairment, cerebellarsymptoms, tremor, apraxia, seizure, grand mal convulsion, palsy, andbalance disorder.

The degree of an adverse effect can, for example, be measured inaccordance with the NCI Common Terminology Criteria for Adverse Eventsv3.0 (CTCAE) (Publish Date: Dec. 12, 2003) in grades. A Grade refers tothe severity of the adverse effects. The CTCAE v3.0 displays grades 1through 5 with unique clinical descriptions of severity for each adverseeffects:

Grade 1: mild adverse effects

Grade 2: Moderate adverse effects

Grade 3: Severe adverse effects

Grade 4: Life-threatening or disabling adverse effects.

Grade 5: Death of the patient.

A “patient” is a human individual who will be or is treated with aCD19×CD3 bispecific antibody. In accordance with the present invention,the patient is suspected/assumed to comprise or already comprisesmalignant CD19 positive lymphocytes (in particular B cells). In thelatter case, said patient has already been diagnosed to comprise suchcells. These malignant CD19 positive lymphocytes (in particular B cells)are present in a patient developing and/or suffering from leukemiaand/or lymphoma. In accordance with the present invention a patient isthus in need of a treatment of malignant CD19 positive lymphocytes.Preferably, a patient who will be or is treated with a CD19×CD3bispecific antibody is (or has been) diagnosed in accordance with themethod of the first aspect of the invention as described herein.

A patient may sometimes be called herein a “high risk patient”, ifhis/her total B cell count is less than 50 B cells per microliter ofperipheral blood as described herein. Similarly, a patient may sometimesbe called herein a “low risk patient”, if his/her total B cell count isgreater than 50 B cells per microliter of peripheral blood as describedherein.

“Mediated by” when used in the context of the method of the first aspectof the present invention means that adverse effects that a patient mayor may not encounter are caused by the administration of a CD19×CD3bispecific antibody. Put it differently, the CD19×CD3 antibody is thecausative agent that may cause potential adverse effects in a patient.

The administration may be in the form of a bolus administration orcontinuous administration, with continuous administration beingpreferred.

In accordance with the present invention by the term “sample” isintended any biological sample obtained from a human patient containingpolynucleotides or polypeptides or portions thereof. Biological samplesinclude body fluids (such as blood, serum, plasma, urine, saliva,synovial fluid and spinal fluid) and tissue sources found to malignantCD19 positive lymphocytes. Methods for obtaining tissue biopsies andbody fluids from patients are well known in the art. Generally, abiological sample which includes peripheral blood mononuclear cells(PBMCs), in particular B cells and T cells is preferred as a source.

A sample which includes peripheral blood mononuclear cells (PBMCs), inparticular B cells and T cells is preferably taken from peripheral bloodof a human patient.

Other preferred samples are whole blood, serum, plasma or synovialfluid, with plasma or serum being most preferred. However, a sample fromperipheral blood of a human patient is particularly preferred.

A “B:T cell ratio” as used herein refers to the ratio of the number of Bcells and the number of T cells. It is preferably determined in a sampletaken from a human patient. Preferably, the sample is taken from theperipheral blood of a human patient. The number of B or T cells, forexample, in a peripheral blood sample can be determined by any meansusually applied in the art, for example, by FACS analysis.

The B:T cell ratio of the patient population treated according to thepresent invention is preferably about 1:5 or lower including a B:T cellratio of about 1:6, 1:7, 1:8, 1:9, 1:10, 1:11, 1:12, 1:13, 1:14, 1:15,1:20, 1:100, 1:200, 1:400, 1:500, 1:1000, 1:2000, 1:3000, 1:4000, 1:5000or even lower, with less than about 1:8, 1:9, 1:10, 1:50, 1:100, 1:500,1:1000 being indicative for a risk of potential adverse effects for saidpatient.

“Determining the B:T cell ratio” includes

-   (a) determining the total B cell number in a sample from a patient,    preferably in a peripheral blood sample of the patient;-   (b) determining the total T cell number in sample from a patient,    preferably in a peripheral blood sample of the patient; and-   (c) calculating the ratio of the B cell number of step (a) and the T    cell number of step (b) in order to obtain a B:T cell ratio.    Of note, a low B:T cell ratio can also be seen as high T:B ratio;    and vice versa. Accordingly, the ratios provided herein for a low    B:T cell ratio would then have to be reversed.

In contrast, patients showing a B:T cell ratio higher than about 1:5including a B:T cell ratio of higher than about 1:4, 1:3, 1:2, 1:1, 2:1,3:1, 4:1, 5:1, 6:1, 7:1, 8:1, 9:1, 10:1 or higher, have a decreased riskof suffering from potential adverse effects upon administration of aCD19×CD3 bispecific antibody. Thus, these patients have higher numbersof B cells as compared to T cells and higher numbers of B cells thancompared to the at risk patients for CNS events.

A “total B cell count” as used herein refers to the number of B cells inthe patient. It is preferably determined in a sample taken from a humanpatient. Preferably, the sample is taken from the peripheral blood of ahuman patient. The number of B cells, for example, in a peripheral bloodsample can be determined by any means usually applied in the art, forexample, by FACS analysis, and provided by the total number of B cellsper microliter of peripheral blood.

The total B cell count in the patient is preferably about 50 Bcells/microliter of peripheral blood or greater, including a total Bcell count of about 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62,63, 64, 65, 66, 67, 68, 69, 70. 71, 72, 73, 74, 75, 76, 77, 78, 79, 80,81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98,99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112,113, 114, 115 or greater per microliter of peripheral blood beingpreferred.

Generally, the preferred minimum number of total B cells shouldpreferably be greater than 50 total B cells, more preferably greaterthan 40, 41, or 42 total B cells or even more preferably greater than 35total B cells per microliter of peripheral blood as indicative for alower risk of adverse effects for a patient when treated with a CD19×CD3bispecific antibody.

Generally, as being indicative for a higher risk of adverse effects fora patient when treated with a CD19×CD3 bispecific antibody, the total Bcell count in the patient is preferably less than 50 total B cells permicroliter of peripheral blood, more preferably less than 42, 41 or 40total B cells per microliter of peripheral blood, even more preferablyless than 35 total B cells per microliter of peripheral blood.

It is apparent that if the cut-off value for high risk patients would beadjusted from, e.g. less than 50 or 40 B cells per microliter ofperipheral blood to less than 35 B cells per microliter of peripheralblood, with less than 35 B cells per microliter of peripheral bloodbeing indicative of a higher risk of adverse effects, then the cut-offvalue for low risk patients is adjusted accordingly, e.g., from greaterthan 50 or 40 B cells per microliter of peripheral blood to greater than35 B cells per microliter of peripheral blood, with greater than 35 Bcell per microliter of peripheral blood being indicative of a lower riskof adverse effects.

Accordingly, the present invention also envisages a method for assessing(analysing) the risk of potential adverse effects for a human patientmediated by the administration of a CD19×CD3 bispecific antibody to saidpatient comprising determining in a sample from said patient the ratioof B cells to T cells of said patient, wherein a ratio of higher thanabout 1:5 or a total B cell count of greater than about 50 Bcells/microliter of peripheral blood is indicative for a decreased riskof potential adverse effects for said patient.

Patients having been assessed to have a decreased risk of potentialadverse effects do not necessarily have to be made subject to the two orthree stage treatment regimen described herein. Accordingly, these lowrisk patients can be treated in the normal way, i.e., there is notnecessarily a need to start with a low dose and increasing doses overtime. These low risk patients can already be treated with a second dose,optionally followed by a third dose as described herein.

However, in case these low risk patients when being monitored duringtherapy may be classified as high risk patients, dependent on the B cellcount as described herein and, thus, may have to be subject to thetreatment regimen as described herein for high risk patients.

Having observed that patients who have a B:T cell ratio of about 1:5 orlower and/or a total B cell count of lower than about 50 Bcells/microliter of peripheral blood are at an increased risk ofpotential adverse effects, the inventors developed a concept that allowsthe treatment of these patients with a CD19×CD3 bispecific antibody.Bearing this in mind, it has been elucidated that the T cells of suchhigh risk patients have to be pre-adapted or partially activated by theadministration of a low dose of antibody for several days before thedose can then be escalated. So it has been found that a significantdecrease in dose given per time unit potentially increases tolerabilityto said antibody in the high risk patients. In essence, the inventorsfound that “adapting” a patient to a CD19×CD3 bispecific antibody priorto the therapy with a CD19×CD3 bispecific antibody is beneficial foravoiding undesired adverse effect (particularly the unwantedneurological reactions) (see Examples 6 and 7).

Accordingly, the present invention relates in a second aspect to amethod (dosage regimen) for administering a CD19×CD3 bispecific antibodyto a human patient having a B:T cell ratio of about 1:5 or lower and/orless than a total B cell count of 50 B cells/microliter of peripheralblood, comprising:

(a) administering a first dose of said antibody for a first period oftime; and consecutively

(b) administering a second dose of said antibody for a second period oftime;

wherein said second dose exceeds said first dose.

In a further embodiment, the present invention relates to a method(dosage regimen or dosage schedule) for treating identified humanpatients at risk for potential adverse effects as a result of treatmentby the administration of a CD19×CD3 bispecific antibody by amelioratingand/or preventing potential adverse effects comprising:

-   -   (a) determining the ratio of B cells to T cells and/or total B        cells in a sample from said patient;    -   (b) identifying human patients at risk for potential adverse        effects based on the results of the determining step; and    -   (c) administering the CD19×CD3 bispecific antibody to the        identified human patients at risk for potential adverse effects        according to a specific dosage schedule to reduce the        possibility of potential adverse effects in said human patients.

The method identifies patients with a total B cell count of less thanabout 50 B cells per microliter of peripheral blood of said patient asindicative for a risk of potential adverse effects for said patient. Oralternatively, the method identifies patients with a ratio of B cells toT cells of about 1:5 or lower in a sample of peripheral blood of saidpatient as indicative for a risk of potential adverse effects for saidpatient.

The method of the present invention provides for the specific dosageschedule for said patient comprises:

(c1) administering a first dose of a CD19×CD3 bispecific antibody for afirst period of time; and consecutively

(c2) administering a second dose of said antibody for the first periodof time or for a second period of time; and optionally consecutively,and

(c3) administering a third dose of said antibody for the first or secondperiod of time or for a third period of time,

wherein said second dose exceeds said first dose and said third doseexceeds said second dose.

The method of the present invention further comprising periodicallydetermining the ratio of B cells to T cells and/or total B cell countsduring administration of said antibody.

Additionally, the method also provides that the specific dosage schedulemay be adjusted based upon the results of periodically determining theratio of B cells to T cells and/or total B cell count to reduce the riskfor potential adverse effects during the treatment.

It will be understood that in the context of the present invention, theterm “method” includes a “dosage regimen” to be used in a method of thepresent invention.

In the context of the present invention “administration of a CD19×CD3bispecific antibody” or “administering a CD19×CD3 bispecific antibody”or any other grammatical form thereof means that the CD19×CD3 antibodyis in the form of a pharmaceutical composition, optionally comprising apharmaceutically acceptable carrier. Accordingly, it is to be understoodthat a pharmaceutical composition comprising a CD19×CD3 bispecificantibody is administered to a human patient.

The term “administering” in all of its grammatical forms meansadministration of a CD19×CD3 bispecific antibody (in the form of apharmaceutical composition) either as the sole therapeutic agent or incombination with another therapeutic agent. It is thus envisaged thatthe pharmaceutical composition of the present invention are employed inco-therapy approaches, i.e. in co-administration with other medicamentsor drugs, for example, other medicaments for treating malignant CD19positive lymphocytes in a patient and/or any other therapeutic agentwhich might be beneficial in the context of the methods of the presentinvention.

For example, if the methods of the invention are carried out for thetreatment of B-lineage acute lymphoblastic leukemia or aggressive NHL,it can advantageously be combined with intrathecal chemotherapy in orderto eliminate target B cells from the CNS. For example, the intrathecalchemotherapy could be performed prior to the administration of theCD19×CD3 bispecific single chain antibody according to the methodsdescribed herein.

The administration of a pharmaceutical composition referred to herein ispreferably an intravenous administration. It follows that in the methodsof the present invention the route of administration in step (a) and/orthe route of administration in step (b) is intravenous. It may beadministered as a bolus injection or continually (continuously), withcontinually being preferred.

The administration of a CD19×CD3 bispecific antibody (for example in theform of a pharmaceutical composition) can be a bolus injection orcontinually or as also sometimes used herein continuously, withcontinually or continuously being preferred. A continual administrationrefers to an administration which is essentially without interruption.“Essentially without interruption” includes a continual administrationusually without an uninterrupted flow or spatial extension.

In some embodiments, said first dose is not therapeutically active, i.e.it is a subtherapeutic dose. Without being strictly bound, for thepurpose of the present invention a dose of 5 μg/m²/d or lower is held tobe subtherapeutic.

In a preferred embodiment of the present invention the second dose istherapeutically active. By “therapeutically effective amount” or“therapeutically active” is meant a dose of a CD19×CD3 bispecificantibody that produces the therapeutic effects for which it isadministered.

The exact dose will depend on the purpose of the treatment, and will beascertainable by one skilled in the art using known techniques. As isknown in the art and described above, adjustments for age, body weight,general health, sex, diet, drug interaction and the severity of thecondition may be necessary, and will be ascertainable with routineexperimentation by those skilled in the art. The therapeutic effect ofthe respective methods or method steps of the present invention isadditionally detectable by all established methods and approaches whichwill indicate a therapeutic effect. It is, for example, envisaged thatthe therapeutic effect is detected by way of surgical resection orbiopsy of an affected tissue/organ which is subsequently analyzed by wayof immunohistochemical (IHC) or comparable immunological techniques.Alternatively it is also envisaged that the tumor markers in the serumof the patient (if present) are detected in order to diagnose whetherthe therapeutic approach is already effective or not. Additionally oralternatively it is also possible to evaluate the general appearance ofthe respective patient (fitness, well-being, decrease of tumor-mediatedailment etc.) which will also aid the skilled practitioner to evaluatewhether a therapeutic effect is already there. The skilled person isaware of numerous other ways which will enable him or her to observe atherapeutic effect of the compounds of the present invention.

In a third aspect, the present invention relates to a method fortreating malignant CD19 positive lymphocytes in a human patient having aB:T cell ratio of about 1:5 or lower and/or less than a total B cellcount of 50 B cells/microliter of peripheral blood, said methodcomprising:

-   (a) administering a first dose of a CD19×CD3 bispecific antibody for    a first period of time; and consecutively-   (b) administering a second dose of said antibody for a second period    of time;    wherein said second dose exceeds said first dose.

Malignant CD19 positive lymphocytes (in particular B cells) are found inleukemia and/or lymphoma. Accordingly, the CD19 positive lymphocytes arein a preferred embodiment lymphoma or leukemia cells.

“Malignant” describes lymphocytes (in particular B cells) thatcontribute to a progressively worsening disease, in particular lymphomaor leukemia and the diseases described herein. The term is most familiaras a description of cancer, here lymphoma and leukemia and the diseasesdescribed herein. Malignant CD19 positive lymphocytes (in particular Bcells) are not self-limited in their growth, are capable of invadinginto adjacent tissues, and may be capable of spreading to distanttissues (metastasizing). Malignant when used herein is synonymous withcancerous.

However, as “normal” (non-malignant) lymphocytes (in particular B cells)also express CD19, it is to be expected that the CD19×CD3 bispecificantibody also binds these normal lymphocytes (in particular B cells) andupon recruiting cytotoxic T cells (because of the second specificity ofthe bispecific CD19xCD13 antibody) depletes these normal B cells. Yet,it is expected that the population of these normal B cells isreconstituted in the absence of the CD19×CD3 bispecific antibody. It wasobserved by Leandro and co-workers that after their depletion by ananti-CD20 antibody, B cells were reconstituted in rheumatoid arthritispatients (Arthritis Rheum. 2006 February: 54(2):613-20). As CD20,likewise CD19 is expressed on almost all B cells, it can be expectedthat B cells upon depletion by the bispecific CD19×CD3 antibody arereconstituted, too.

The lymphoma is preferably indolent or aggressive B cell non-Hodgkinlymphoma (B NHL), mantle cell lymphoma (MCL) or chronic lymphaticleukemia (CLL). Within the meaning of the invention, the term “B cellnon-Hodgkin lymphoma” or “B cell derived non-Hodgkin lymphoma” comprisesboth indolent and aggressive B cell non-Hodgkin lymphoma (B NHL). Theterm “indolent or aggressive B cell non-Hodgkin lymphoma (B NHL)” asused herein represents malignant B cell-derived tumorous diseases.Indolent B NHL are low malignant lymphomas. Aggressive B-NHL are highmalignant lymphomas. The B cell non-Hodgkin lymphoma (B NHL) mayadvantageously be a follicular lymphoma, lymphoplasmacytic lymphoma,marginal zone cell lymphoma, mantle cell lymphoma (MCL), diffuse large Bcell lymphoma (DLBCL), Burkitt's lymphoma, small lymphocytic lymphoma(SLL/CLL) and any other B cell derived subtype. The term “B cellleukemia” as used herein may advantageously be any B cell leukaemia(e.g. chronic lymphocytic leukaemia or acute lymphocytic leukaemia). Forfurther reference see e.g. http://www.cancer.org. Preferably, indolentnon-Hodgkin B cell lymphoma may be treated with a bispecific singlechain antibody directed against both human CD3 and human CD19 asdemonstrated in the following examples.

The leukemia is preferably B-lineage acute lymphoblastic leukemia (ALL).

In a fourth aspect, the present invention relates to a method forameliorating and/or preventing an adverse effect mediated by theadministration of a CD19×CD3 bispecific antibody to a human patienthaving a B:T cell ratio of about 1:5 or lower and/or less than a total Bcell count of 50 B cells/microliter of peripheral blood, said methodcomprising:

(a) administering a first dose of said antibody for a first period oftime, and consecutively

(b) administering a second dose of said antibody for a second period oftime;

wherein said second dose exceeds said first dose.

The adverse effect is preferably a neurological reaction, preferably oneor more selected from the group consisting of: confusion, ataxia,disorientation, dysphasia, aphasia, speech impairment, cerebellarsymptoms, tremor, apraxia, seizure, grand mal convulsion, palsy, andbalance disorder (see also Examples 2 and 3).

Specifically, neurological reactions observed during the starting phaseof treatment with the CD19×CD3 bispecific antibody include for exampleconfusion and disorientation. “Confusion” as used herein refers to lossof orientation which is the ability to place oneself correctly in theworld by time, location, and personal identity, and often memory whichis the ability to correctly recall previous events or learn newmaterial. The patients usually have difficulties to concentrate andthinking is not only blurred and unclear but often significantly sloweddown. Patients with neurological reactions also suffer from loss ofmemory. Frequently, the confusion leads to the loss of ability torecognize people and/or places, or to tell time and the date. Feelingsof disorientation are common in confusion, and the decision-makingability is impaired. Neurological reactions further comprise blurredspeech and/or word finding difficulties. This disorder may impair both,the expression and understanding of language as well as reading andwriting. Besides urinary incontinence, also vertigo and dizziness mayaccompany neurological reactions in some patients.

The occurrence of neurological reactions in the treatment of B celldependent lymphatic or leukemic malignancies with the CD19×CD3bispecific antibody may be further influenced by the following factors:

1. Presence of Drug

The CD19×CD3 bispecific antibody retargets T cell cytotoxicity tomalignant CD19 positive lymphocytes present, for example, in B celllymphoma or leukemia cells. In light of this, it can be reasonablyassumed that it is the presence of CD19×CD3 bispecific antibody in thebody of a patient which is responsible for the adverse effects.Furthermore, side effects are observed only in parts of the body wherethe CD19×CD3 bispecific antibody is biologically active. Accordingly,neurological reactions upon treatment with the CD19×CD3 bispecificantibody are assumed to be dependent on the presence of said antibody inthe cerebrospinal fluid (CSF; liquor) of the patient. This may besupported by the fact that the CD19×CD3 bispecific antibody as well as Tcells have only been found in the CSF of NHL patients with a low B:Tcell ratio. As explained herein, this patient population has anincreased risk for the development of neurological reactions uponantibody treatment. This finding may suggest that the CD19×CD3bispecific antibody is able to enter the perivascular space dividing theblood vessels and the CNS (including the brain), in high risk NHL andALL patients. There, the CD19×CD3 bispecific antibody may then engage Tcells to target local B cells (either benign or malign) which possiblyleads to local cytokine release which in turn could cause neurologicalreactions.

2. Drug Dose

Further, the neurological reactions seem to be dependent on the dose ofthe CD19×CD3 bispecific antibody. For example, neurological reactionshave not been observed upon continuous administration of 5 μg/m² bodysurface area of CD19×CD3 bispecific antibody, but with 15 μg/m² bodysurface area or more CD19×CD3 bispecific antibody in the high risk groupof patients. For this reason, as mentioned herein, a dose of less than 5μg/d/m² is deemed to be subtherapeutic. The effect of the drug dose isevident from the data shown in the appended examples. This observationmay imply a dose dependency of neurological reactions in high riskpatients with low B:T cell ratio and/or low total B cell counts.

3. Presence of Target Cells and Effector Cells

As set forth above, the neurological reactions upon CD19×CD3 bispecificantibody-treatment are assumed to depend on the presence of i) targetcells, i.e. CD19-antigen carrying B cells and ii) effector cells, i.e.cytotoxically active T cells carrying the CD3 antigen, in the PVS/CNS.

In view of this, it is intriguing to hypothesize that the depletion ofe.g. the target B cell from the PVS/CNS should result in the avoidanceof neurological reactions. In fact, this is exactly what has beenobserved in the mentioned phase II study in which B lineage acutelymphoblastic leukemia (ALL) patients are being currently treated withthe CD19×CD3 bispecific antibody:

In ALL, there is generally a high incidence of leukemic lesions in theCNS. Therefore, each of the ALL patients enrolled in the clinical phaseII study referred to herein had received standard ALL therapies in thepast, including methotrexate i.v. and/or intrathecal chemotherapy, inorder to prevent central nervous system relapses. Some of them receivedin addition irradiation of the neuroaxis. The ALL patients thereafterreceived a consolidation therapy, i.e. they obtained several fourweek-treatment cycles of continuous administration of 15 μg/d/m² ofCD19×CD3 bispecific antibody. Only one of the thus far enrolled 17 ALLpatients who have been treated with the CD19×CD3 bispecific antibody hasdeveloped neurological reactions. This patient was one out of 11patients belonging to the high risk group having a B:T cell ratio lowerthan 1:5. None of the six patients of the low risk group with a B:T cellratio higher than 1:5 showed neurological reactions. It is thereforehypothesized that the mentioned (pre-symptomatic) central nervous system(CNS) treatment reduced the risk of a neurological reaction in the ALLpatient in that the B lymphocytic target cells have been removed fromthe PVS and CNS, including the brain. However, in the absence of Btarget cells in these tissues, there is no full activation of thecytotoxic T cells. Therefore, less frequent neurological reactions couldbe observed in said patient populations (see also FIGS. 2A and 2B)

Accordingly, the absence of one of the above factors, in the mentionedcase the presence of target B cells in the PVS/CNS, could possibly helpto prevent neurological reactions. However, for example, intrathecalchemotherapy is not the therapy of choice in NHL treatment. Forinstance, it is not effective in indolent NHL therapy, and it is not yetknown whether it could be a treatment option for aggressive NHL. Inaddition, intrathecal chemotherapy is highly toxic for ALL patients andtherefore associated with considerable health risks.

In light of the above, the depletion of any one of the above indicatedfactors without losing therapeutic efficacy is no trivial task since itis for example not easily possible to avoid the presence of B cells inthe PVS/CNS of NHL. Furthermore, it has also been found that othermeasures, including the pre- or co-administration of high doses ofsteroids could not prevent neurological reactions in the high riskpatients.

However, by way of applying the methods/dosage regimens of the presentinvention, it is possible to ameliorate and/or prevent adverse effectsfor patients who are at an increased risk of such adverse effects ifthey have a B:T cell ratio of about 1:5 or lower and/or a total B cellcount of less than 50 B cells/microliter of peripheral blood. Thepresent invention envisages providing dosage regimens (methods) whichare even independent of the above mentioned factors that could influencea treatment with a CD19×CD3 bispecific antibody.

Thus, the present invention in a preferred aspect relates to a methodfor assessing (analysing) the risk of potential adverse effects for ahuman patient mediated by the administration of a CD19×CD3 bispecificantibody to said patient comprising determining the ratio of B cells toT cells in a sample from said patient, wherein a ratio of about 1:5 orlower or determining a total B cell count in a sample from a patient,wherein a total B cell count of less than 50 B cells/microliter ofperipheral blood, is indicative for a risk of potential adverse effectsfor said patient, wherein said patient is

(a) administered a first dose of said antibody for a first period oftime; and is consecutively

(b) administered a second dose of said antibody for a second period oftime;

wherein said second dose exceeds said first dose;

for

-   (i) treating malignant CD19 positive lymphocytes; and/or-   (ii) for ameliorating and/or preventing an adverse effect mediated    by the administration of a CD19×CD3 bispecific antibody.

Preferably, in this preferred aspect, the patient is administered athird dose of said antibody for a third period of time as described inherein below. Accordingly, the embodiments and aspects described hereinin the context of the three-stage method (dose regimen) are applicableto this preferred aspect.

In one aspect of the methods of the present invention said second periodof time exceeds said first period of time. The term “exceeds” means thatthe second period of time is at least one day longer than the firstperiod of time.

Each of the methods (dosage regimens) of the present invention can berepeated, for example, for one, two, three, four, five, six, or moretimes and in any event as often as there is a beneficial effect for apatient in ameliorating and/or treating malignant CD19 positivelymphocytes, thereby treating lymphoma or leukemia. Dependent on theratio of the B:T cell ratio and/or total B cell count of a patient, inaccordance with the teaching of the present invention, the practitionercan decide as to whether the patient has to be “adapted” to a furthertreatment with a CD19×CD3 bispecific antibody prior by applying thedosage regimens of the present invention (i.e., administering a low doseof a CD19×CD3 bispecific antibody prior to administering a higher dosein order to “adapt” the patient).

In one embodiment, the dosage of the bispecific antibody can becalculated. For example based upon the data showing the importance ofthe B:T cell ratio and/or total B cell count in determining theappropriate dose of the CD19×CD3 bispecific antibody in the populationof patients at risk for CNS events, a calculation or formula may be usedto assist the practitioner in determining appropriate dosages for thispatient population. This formula is based on the relationship of theCD19×CD3 bispecific antibody (blinatumomab) dose and the B and T cellcounts. For example,

${{c\mspace{14mu}\left( {{contant}\mspace{14mu}{factor}} \right)} = \frac{C\; D\; 19 \times C\; D\; 3\mspace{14mu}{bispecific}\mspace{14mu}{antibody}\mspace{14mu}{dose} \times T\mspace{14mu}{cell}\mspace{14mu}{count}}{B\mspace{14mu}{cell}\mspace{14mu}{count}}};$meaning that

${C\; D\; 19 \times C\; D\; 3\mspace{14mu}{bispecific}\mspace{14mu}{antibody\_ dose}} = \frac{c \times B\mspace{14mu}{cell}\mspace{14mu}{count}}{T\mspace{14mu}{cell}\mspace{14mu}{count}}$or alternatively,individualized doses based upon individual patient's B:T cell ratio ortotal B cell counts can be utilized, where the bispecific antibody doseis calculated by multiplying either the B:T cell ratio or total B cellcount by the Constant factor.

It must be understood that the dose or day ranges given herein areillustrated by increments of one, two, three, four or five. Theseranges, however, in case of increments higher than one also encompasssmaller increments, for example those exemplified by increments of one(10 to 30 includes for example 10, 11, 12, 13, 13 etc. up to 30), orstill smaller increments, for example values after the decimal point.

In another aspect of the present invention, it is envisaged that saidfirst period of time is at least 3 days long, whereby even longerperiods of time of for example 8, 9, 10, 11, 12, 13 or 14 days are notexcluded. “Longer” is thereby not limited to a (one) complete day as thelowest time unit, i.e. ½ days, or fully hours are also conceivable. Itis however preferred that the smallest time unit is one full day.Accordingly, said first period of time exceeds 3 days. More preferably,it is envisaged that said first period of time is between 3 days and 10days, with 7 days being particularly preferred.

As used herein, a time interval which is defined as “X to Y” equateswith a time interval which is defined as “between X and Y”. Both timeintervals specifically include the upper limit and also the lower limit.This means that for example a time interval “3 to 10 days” or between “3to 10 days” includes a period of time of one, two, three, four, five,six, seven and/or eight days.

As mentioned herein, the inventors observed that “adapting” a humanpatient having a B:T cell ratio of about 1:5 or lower to the treatmentwith a CD19×CD3 bispecific antibody during a first period of time allowsthe treatment of the human patient with an increased second dose of theantibody for a second period of time, whereby adverse effects (inparticular neurological reactions) can be better controlled, i.e., couldbe avoided or at least kept within an acceptable grade in accordancewith the CTCAE.

However, for achieving this improvement it is required to “adapt” thehuman patient having a B:T cell ratio of about 1:5 or lower and/or atotal B cell count of less than 50 B cells/microliter of peripheralblood to the CD19×CD3 bispecific antibody by administering a first doseof the antibody for a first period of time (wherein said first dose islower than the consecutive (second) dose). The administration can be abolus injection or a continuous administration, whereby a continuousadministration is preferred.

Likewise the duration of the first period of time, the duration of thesecond period of time may be variable in view of, for example, the age,sex, body weight, etc. of the human patient.

Accordingly, in another aspect of the present invention, it is envisagedthat said second period of time is at least 18 days long, whereby evenlonger periods of time of for example 19, 20, 25, 30, 35, 40, 45, 49,50, 55, 60, 65, 60, 65, 70, 75, 80, 81, 82, 83, 84, 85, 86, 87, 88 or 90days are not excluded. “Longer” is thereby not limited to a (one)complete day as the lowest time unit, i.e. ½ days, or fully hours arealso conceivable. It is however preferred that the smallest time unit isone full day.

Accordingly, said second period of time exceeds 18 days. Morepreferably, it is envisaged that said second period of time is between18 days and 81 days, with 21 or 49 days being particularly preferred.

As used herein, a time interval which is defined as “X to Y” equateswith a time interval which is defined as “between X and Y”. Both timeintervals specifically include the upper limit and also the lower limit.This means that for example a time interval “18 to 81 days” or between“18 to 81 days” includes a period of time of 1, 2, 3, 4, 5, 6, 7, 8, 9,10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27,28, 29, 30, 32, 34, 36, 38, 40, 42, 44, 46, 48, 50, 52, 54, 56, 58, 60,61, 62, 63 and/or 64 days.

In a more preferred embodiment of the methods/dosage regimens of thepresent invention, said first period of time is between 3 days and 10days, and said second period of time is between 18 and 81 days.

In an even more preferred embodiment, said first period of time is 7days and said second period of time is 21 or 49 days.

In the clinical trials mentioned herein, it was observed that a dose of15 μg/m²/d in the treatment of NHL effected tumor shrinkage as could bevisualized in computer tomography. It was also observed that a dose of15 μg/m²/d in the treatment of ALL resulted in minimal residual diseaseand could even eliminate MRD.

Minimal residual disease (MRD) is the name given, to small numbers ofleukemic/lymphoma cells that remain in the patient during treatment orafter treatment when the patient is in remission (no symptoms or signsof disease). Up until a decade ago none of the tests used toassess/detect cancer, were sensitive enough to detect MRD. Now, however,very sensitive molecular biology tests are available—based on DNA, RNAor Proteins—and these can measure minute levels of cancer cells intissue samples, sometimes as low as 1 cancer cell in million normalcells.

In cancer treatment, particularly leukemia, MRD testing has severalimportant roles: determining whether treatment has eradicated the canceror whether traces remain, comparing the efficacy of differenttreatments, monitoring patient remission status and recurrence of theleukemia or cancer and choosing the treatment that will best meet thoseneeds (personalization of treatment)

Accordingly, in a further aspect of the methods/dosage regimens of thepresent invention, said first dose is between 1 and 15 μg/m²/d, i.e. 1,2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15 μg/m²/d. Particularlypreferred is a dose of 5 or 15 μg/m²/d.

As used herein, a dose interval which is defined as “between X and Y”equates with a dose interval which is defined as “X to Y”. Both doseintervals specifically include the upper limit and also the lower limit.This means that for example a dose interval “between 1 and 15” or “1 to15” includes a dose of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15μg/m²/d. “d” denotes one day.

“m²” denotes a square meter of a patient's body surface (BSA). The“normal” average BSA is generally taken to be about 1.73 m² for anadult, for a neonate it is about 0.25 m², for a 2 year old child it isabout 0.5 m², for a 9 year old child it is about 1.07 m², for a 10 yearold child it is about 1.14 m², for a 12-13 year old child it is about1.33 m², for men it is about 1.9 m² and for women it is about 1.6 m².

However, the BSA can also be calculated more precisely by one of thefollowing formulas (each of these formulas can be applied whencalculating the BSA):

The Mosteller formula (Mosteller, N Engl J Med 1987 Oct. 22; 317(17):1098):BSA (m²)=([Height (cm)×Weight (kg)]/3600)^(1/2) or in inches and pounds:BSA (m²)=([Height (in)×Weight (lbs)]/3131)^(1/2)

The DuBois formula (DuBois, Arch Int Med 1916 17:863-871):BSA (m²)=0.007184×Height (cm)^(0.725)×Weight (kg)^(0.425)

The Haycock formula (Haycock, The Journal of Pediatrics 1978 93:1:62-66):BSA (m²)=0.024265×Height (cm)^(0.3964)×Weight (kg)^(0.5378)

The Gehan formula (Gehan, Cancer Chemother Rep 1970 54:225-35):BSA (m²)=0.0235×Height (cm)^(0.42246)×Weight (kg)^(0.51456)

The Boyd formula (Boyd, University of Minnesota Press, 1935)BSA (m²)=0.0003207×Height (cm)^(0.3)×Weight(grams)^((0.7285-(0.0188×log 10(grams))

It is generally preferred that each of the doses disclosed herein can beconverted from amount in μg/m²/d into μg/d by multiplying the respectivedose with the factor 1.9. Accordingly, each of the doses disclosedherein can be applied in the methods and uses by multiplying it with thefactor 1.9. For example, a dose of 5 μg/m²/d is converted into 9.5 μg/d,a dose of 15 μg/m²/d is converted into 28.5 μg/d and a dose of 60μg/m²/d is converted into 114 μg/d. It is preferred that a decimal digitthat results from the multiplication is either rounded up or roundeddown, respectively, to a whole number. For example, a dose of 9.5 μg/dcan be rounded down to 9 μg/d and a dose of 28.5 μg/d can be roundeddown to 28 μg/d, and after rounding the 60 μg/m²/d dose, it can also beconverted to e.g. 112 μg/d (4×28 μg/d). Likewise, a dose of 9.5 μg/d canbe rounded up to 10 μg/d and a dose of 28.5 μg/d can be rounded up to 29μg/d. One skilled in the art can convert the dosing from amount μg/m²/dinto μg/d and rounding up or down to obtain an equivalent dose using thefactor of 1.9.

The term “μg” [microgram] includes “μg of the CD19×CD3 bispecificantibody preparation”. It is preferred that not more than 10% of saidCD19×CD3 bispecific antibody preparation is incorrectly folded. Itfollows that in a preferred embodiment, 90%, 91%, 92%, 93%, 94%, 95%,96%, 97%, 98%, 99% or even 100% of the CD19×CD3 bispecific antibody iscorrectly folded. It is also conceivable that the antibody preparationmay optionally comprise further ingredients, for example alyoprotectant, a surfactant, a filler, a binder, and/or bulking agentetc. The amount of such further ingredients is, preferably, not includedin the term “μg” as used in the context of the “dose” and/or methods(dosage regimens) of the present invention.

A dose of, for example, 1 μg/m²/d means that 1 μg of the CD19×CD3bispecific antibody is administered evenly or continuously across oneday per square meter body surface. “Continuously across one day” refersto an infusion which is allowed to proceed permanently withoutinterruption.

In a further aspect of the methods/dosage regimen of the presentinvention, said second dose is between 15 and 60 or 15 and 90 μg/m²/d,i.e. 15, 20, 25, 30, 35, 40, 45, 50, 55 and 60 μg/m²/d or 15, 20, 25,30, 35, 40, 45, 50, 55, 60, 70, 80 and 90 μg/m²/d. Particularlypreferred is a dose of 60 or 90 μg/m²/d. Said second dose is thustherapeutically active. In a preferred embodiment, said first dose isbetween 5 and 15 μg/m²/d and said second dose is between 15 and 60 or 15and 90 μg/m²/d.

As used herein, a dose interval which is defined as “between X and Y”equates with a dose interval which is defined as “X to Y”. Both doseintervals specifically include the upper limit and also the lower limit.This means that for example a dose interval “between 15 and 60” or “15to 60” includes a dose of 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25,26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43,44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59 and/or 60μg/m²/d.

It must be understood that the ranges given herein are illustrated byincrements of five. These ranges, however, also encompass smallerincrements, for example those exemplified by increments of one (10 to 30includes for example 10, 11, 12, 13, 13 etc. up to 30), or still smallerincrements, for example values after the decimal point.

In a further aspect to reduce the risk of the possibility of adverseeffects, particularly CNS adverse effects, the methods/dosage regimen orschedule should be administered with the intent to reduce the initialtreatment intensity of the CD19×CD3 bispecific antibody to the patient.For example, an approach to reducing the initial treatment would includea step up dosage regimen or schedule to “adapt” the patient, beginningwith an initial dosage of the CD19×CD3 bispecific antibody of between 5and 15 μg/m²/d [or between 9-10 μg/d and 28-29 μg/d] with a further stepup in a second dose of 30, 45 or 60 μg/m²/d [57 μg/d, 85-86 μg/d or112-116 μg/d, respectively] in gradual steps. Flating dosing can beutilized for each of these dosages using the conversion factor of 1.9and rounding up or down. Also dosage regimens of 5 μg/m²/d to 15 μg/m²/dinitially to 45 μg/m²/d then to 60 μg/m²/d either gradually, or in stepthat could jump some of the intermediate dosage levels if no adverseside effects were observed. Further, dosage regimens or schedules couldinclude administration of 5 μg/m²/d that is increased to 15 μg/m²/d andthen to 60 μg/m²/d or as 5 μg/m²/d or 15 μg/m²/d dosage regimens orschedules that are increased to 60 μg/m²/d over time. The practitionercan increase the dosage of the bispecific antibody if the B:T cellratios and/or total B cell counts continue to remain in the low riskcategory for adverse effects (a B:T cell ration of 1:5 or lower and/or atotal B cell count of less than 50 B cells per microliter of peripheralblood), and particularly CNS adverse effects. But the practitionershould also consider the proper dosage regiment to obtain efficaciousresults for the disease being treated by the CD19×CD3 bispecificantibody, and balance the efficacy of the antibody with the reduction inrisk of adverse effects.

Preferably, not included in the methods for administering a CD19×CD3bispecific antibody, for treating malignant CD19 positive lymphocytes,or for ameliorating and/or preventing an adverse effect mediated by theadministration of a CD19×CD3 bispecific antibody are the followingadministration schemes:

-   (i) 5 μg/m² of the bispecific antibody for one day followed by    administration of 15 μg/m² as daily dose for the remaining period    (second and each further consecutive day); and/or-   (ii) 15 μg/m² of the bispecific antibody for one day followed by    administration of 45 μg/m² as daily dose for the remaining period    (second and each further consecutive day); and/or-   (iii) 5 μg/m² of the bispecific antibody for one day followed by    administration of 15 μg/m² for one day, followed by administration    of 45 μg/m² as daily dose for the remaining period (third and each    further consecutive day); and/or-   (iv) less than 10-80 μg/m² of the bispecific antibody for one day    followed by administration of a dose of 10-80 μg/m² (second and each    further consecutive day); and/or-   (v) less than 10-80 μg/m² of the bispecific antibody for one day    followed by administration of a dose of less 10-80 μg/m² for one    day, followed by administration of a dose of less 10-80 μg/m² (third    and each further consecutive day).    Again flat dosing per day may be used.

As mentioned herein, patients having a B:T cell ratio higher than 1:5and/or a total B cell count of equal to or greater than 50 Bcells/microliter of peripheral blood, do not necessarily have to beadapted to the treatment with a CD19XCD3 bispecific antibody by way ofthe dosage regimen of the present invention. These patients having adecreased risk of potential adverse effects could be treated byadministration of a CD19×CD3 bispecific antibody in a constant dose of 5μg to 75 μg per square meter body surface area per day for at least fourweeks. The administration is preferably a continuous administration.

In another embodiment of the methods (dosage regimen) of the presentapplication, said methods further comprise administering after a firstand second dose for a first and second period of time a third dose ofsaid antibody for a third period of time. Accordingly, the presentinvention provides a three-stage method (dosage regimen).

The administration of said third dose is intravenously. It can beadministered in the form of a bolus injection or continuously, withcontinuously being preferred.

In one aspect of the methods of the present invention said third periodof time exceeds said first and second period of time. The term “exceeds”means that the third period of time is at least one day longer than thefirst and second period of time.

Likewise the duration of the first and second period of time, theduration of the third period of time may be variable in view of, forexample, the age, sex, body weight, etc. of the human patient.

In the three-stage dosage regimen aspect of the present invention, it isenvisaged that said first period of time is at least 3 days long,whereby even longer periods of time of for example 8, 9, 10, 11, 12, 13or 14 days are not excluded. “Longer” is thereby not limited to a (one)complete day as the lowest time unit, i.e. IA days, or fully hours arealso conceivable. It is however preferred that the smallest time unit isone full day.

Accordingly, said first period of time exceeds 3 days. More preferably,it is envisaged that said first period of time is between 3 days and 10days, with 7 days being particularly preferred.

As used herein, a time interval which is defined as “X to Y” equateswith a time interval which is defined as “between X and Y”. Both timeintervals specifically include the upper limit and also the lower limit.This means that for example a time interval “3 to 10 days” or between “3to 10 days” includes a period of time of one, two, three, four, five,six, seven and/or eight days.

In the three-stage dosage regimen aspect of the present invention, it isenvisaged that said second period of time is at least 3 days long,whereby even longer periods of time of for example 8, 9, 10, 11, 12, 13or 14 days are not excluded. “Longer” is thereby not limited to a (one)complete day as the lowest time unit, i.e. ½ days, or fully hours arealso conceivable. It is however preferred that the smallest time unit isone full day.

Accordingly, said first period of time exceeds 3 days. More preferably,it is envisaged that said first period of time is between 3 days and 10days, with 7 days being particularly preferred.

As used herein, a time interval which is defined as “X to Y” equateswith a time interval which is defined as “between X and Y”. Both timeintervals specifically include the upper limit and also the lower limit.This means that for example a time interval “3 to 10 days” or between “3to 10 days” includes a period of time of one, two, three, four, five,six, seven and/or eight days.

In the three-stage dosage regimen aspect of the present invention, it isenvisaged that said third period of time is at least 8 days long,whereby even longer periods of time of for example 8, 9, 10, 11, 12, 13,14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 32,34, 36, 38, 40, 42, 44, 46, 48, 50, 52, 54, 56, 58, 60, 61, 62, 63, 64,65, 66, 67, 68, 69, 70 and/or 71 days are not excluded. “Longer” isthereby not limited to a (one) complete day as the lowest time unit,i.e. ½ days, or fully hours are also conceivable. It is howeverpreferred that the smallest time unit is one full day.

Accordingly, said first period of time exceeds 8 days. More preferably,it is envisaged that said first period of time is between 8 days and 78days, with 14 or 42 days being particularly preferred.

As used herein, a time interval which is defined as “X to Y” equateswith a time interval which is defined as “between X and Y”. Both timeintervals specifically include the upper limit and also the lower limit.This means that for example a time interval “18 to 78 days” or between“18 to 78 days” includes a period of time of 1, 2, 3, 4, 5, 6, 7, 8, 9,10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27,28, 29, 30, 32, 34, 36, 38, 40, 42, 44, 46, 48, 50, 52, 54, 56, 58, 60,61, 62 63, 64, 65, 66, 67, 68, 69, 70 and/or 71 days.

In a more preferred embodiment of the three-stage methods/dosageregimens of the present invention, said first period of time is between3 days and 10 days, and said second period of time is between 3 days and10 days, and said third period of time is between 8 days and 78 days.

In an even more preferred embodiment, said first period of time is 7days, said second period of time is 7 days, and said third period oftime is 14 or 42 days.

In an embodiment of the three-stage methods/dosage regimens of thepresent invention, said third dose exceeds said first and second dose.Said second and third dose are preferably therapeutically active. Ofnote, said second dose exceeds said first dose.

Accordingly, in a further aspect of the three-stage methods/dosageregimens of the present invention, said first dose is between 1 and 15μg/m²/d, preferably between 5 and 15 μg/m²/d, i.e. 1, 2, 3, 4, 5, 6, 7,8, 9, 10, 11, 12, 13, 14, 15 μg/m²/d. Particularly preferred is a doseof 5 or 10 μg/m²/d.

In a further aspect of the three-stage methods/dosage regimens of thepresent invention, said second dose is between 1 and 15 μg/m²/d,preferably between 5 and 15 μg/m2/d, i.e. 1, 2, 3, 4, 5, 6, 7, 8, 9, 10,11, 12, 13, 14, 15 μg/m²/d. Particularly preferred is a dose of 15μg/m²/d.

As used herein, a dose interval which is defined as “between X and Y”equates with a dose interval which is defined as “X to Y”. Both doseintervals specifically include the upper limit and also the lower limit.This means that for example a dose interval “between 1 and 15” or “1 to15” includes a dose of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15μg/m²/d.

In a further aspect of the three-stage methods/dosage regimen of thepresent invention, said third dose is between 15 and 60 μg/m²/d, morepreferably between 20 and 60 μg/m²/d, i.e. 15, 20, 25, 30, 35, 40, 45,50, 55 and 60 μg/m²/d. Particularly preferred is a dose of 60 μg/m²/d.Alternatively, said third dose is between 15 and 90 μg/m²/d, morepreferably between 60 and 90 μg/m²/d, i.e., 15, 20, 25, 30, 35, 40, 45,50, 55, 60, 70, 80 and 90 μg/m²/d.

In a preferred embodiment of the three-stage methods/dosage regimen ofthe present invention, said first dose is between 1 and 15 μg/m²/d, saidsecond dose is between 1 and 15 μg/m²/d, and said third dose is between15 and 60 or 15 and 90 μg/m²/d.

Particularly preferred, said first dose is 5 μg/m²/d, said second doseis 15 μg/m²/d, and said third dose is 60 μg/m²/d. Alternatively, saidthird dose may be 90 μg/m²/d.

As used herein, a dose interval which is defined as “between X and Y”equates with a dose interval which is defined as “X to Y”. Both doseintervals specifically include the upper limit and also the lower limit.This means that for example a dose interval “between 15 and 60” or “15to 60” includes a dose of 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25,26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43,44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59 and/or 60μg/m²/d. Similarly, this means that for example a dose interval “between15 and 90” or “15 to 90” includes a dose of 15, 16, 17, 18, 19, 20, 21,22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39,40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57,58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75,76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 86, 87, 88, 89 or 90μg/m²/d.

In view of the observations made by the present inventors that athree-stage (step) method/dosage regimen aids in avoiding adverseeffects as described herein, the present invention relates to a methodof treating malignant CD19 positive lymphocytes in a human patient, saidmethod comprising (a) administering a first dose of said antibody for afirst period of time; (b) administering a second dose of said antibodyfor a second period of time; and consecutively (c) administering a thirddose of said antibody for a third period of time.

Also, the present invention relates to a method for treating malignantCD19 positive lymphocytes in a human patient, said method comprising (a)administering a first dose of said antibody for a first period of time;(b) administering a second dose of said antibody for a second period oftime; and consecutively (c) administering a third dose of said antibodyfor a third period of time.

Furthermore, the present invention relates to a method for amelioratingand/or preventing an adverse effect mediated by the administration of aCD19×CD3 bispecific antibody to a human patient, said method comprising(a) administering a first dose of said antibody for a first period oftime; (b) administering a second dose of said antibody for a secondperiod of time; and consecutively (c) administering a third dose of saidantibody for a third period of time.

Preferably, the first, second and third period of time are as describedelsewhere herein.

Regarding the doses, it is preferred that the second dose exceeds thefirst dose and the third dose exceeds the second dose as describedelsewhere herein. More preferably, the first dose is 5 μg/m²/d, thesecond dose is 15 μg/m²/d and the third dose is 60 μg/m²/d.Alternatively, the third dose may also be 90 or 120 μg/m²/d. Theequivalent flat doses per day of these doses can be determined by thefactor discussed earlier. For example, a dose of 5 μg/m²/d is convertedinto 9.5 μg/d and rounded to 9 or 10 μg/d, a dose of 15 μg/m² isconverted into 28.5 μg/m²/and rounded to 28 or 29 μg/d, and a dose of 60μg/m²/is converted into 112 μg/m², 114 μg/m² or 116 μg/m² depending uponrounding.

As noted herein above, the present invention relates to methods oftreatment/dosage regimen which employ CD19×CD3 bispecific antibodies,comprising a first binding domain capable of binding to an epitope ofhuman CD3 epsilon chain and a second binding domain capable of bindingto human CD19. Examples for bispecific molecules according to themethods of the invention are described in great detail in WO 99/54440and WO 2004/106381 and WO 2008/119565. All the specific CD19×CD3bispecific antibodies disclosed therein, including their variants,fragments, equivalents etc. are particularly preferred CD19×CD3bispecific antibodies of the present invention.

As used herein, a “CD19×CD3 bispecific antibody” (including a CD19×CD3bispecific single chain antibody) denotes a binding entity (“binder”)having at least one polypeptide chain comprising two binding domainssuch as Dual-Affinity Re-Targeting (DART) antibodies, diabodies, domainantibodies (dAbs) or nanobodies. A particularly preferred CD19×CD3bispecific antibody comprises a single polypeptide chain comprising twobinding domains. Such single chain antibodies are preferred in thecontext of the methods/dosage regimen of the present invention. Eachbinding domain comprises at least one variable region from an antibodyheavy chain (“VH or H region”), wherein the VH region of the firstbinding domain specifically binds to the CD3 epsilon molecule, and theVH region of the second binding domain specifically binds to CD19. Thetwo binding domains are optionally linked to one another by a shortpolypeptide spacer. A non-limiting example for a polypeptide spacer isGly-Gly-Gly-Gly-Ser (G-G-G-G-S) (SEQ ID NO: 23) and repeats thereof.Each binding domain may additionally comprise one variable region froman antibody light chain (“VL or L region”), the VH region and VL regionwithin each of the first and second binding domains being linked to oneanother via a polypeptide linker, for example of the type disclosed andclaimed in EP 623679 B 1, but in any case long enough to allow the VHregion and VL region of the first binding domain and the VH region andVL region of the second binding domain to pair with one another suchthat, together, they are able to specifically bind to the respectivefirst and second binding domains. Such CD19CD3 bispecific single chainantibodies are described in great detail in WO 99/54440 and WO2004/106381.

The term “binding domain” characterizes in connection with the presentinvention a domain of a polypeptide which specifically bindsto/interacts with a given target structure/antigen/epitope. Thus, thebinding domain is an “antigen-interaction-site”. The term“antigen-interaction-site” defines, in accordance with the presentinvention, a motif of a polypeptide, which is able to specificallyinteract with a specific antigen or a specific group of antigens, e.g.the identical antigen in different species. Said binding/interaction isalso understood to define a “specific recognition”. The term“specifically recognizing” means in accordance with this invention thatthe antibody molecule is capable of specifically interacting with and/orbinding to at least two, preferably at least three, more preferably atleast four amino acids of an antigen, e.g. the human CD3 antigen asdefined herein. Such binding may be exemplified by the specificity of a“lock-and-key-principle”. Thus, specific motifs in the amino acidsequence of the binding domain and the antigen bind to each other as aresult of their primary, secondary or tertiary structure as well as theresult of secondary modifications of said structure. The specificinteraction of the antigen-interaction-site with its specific antigenmay result as well in a simple binding of said site to the antigen.Moreover, the specific interaction of the bindingdomain/antigen-interaction-site with its specific antigen mayalternatively result in the initiation of a signal, e.g. due to theinduction of a change of the conformation of the antigen, anoligomerization of the antigen, etc. A preferred example of a bindingdomain in line with the present invention is an antibody. The bindingdomain may be a monoclonal or polyclonal antibody or derived from amonoclonal or polyclonal antibody.

The term “antibody” comprises derivatives or functional fragmentsthereof which still retain the binding specificity. Techniques for theproduction of antibodies are well known in the art and described, e.g.in Harlow and Lane “Antibodies, A Laboratory Manual”, Cold Spring HarborLaboratory Press, 1988 and Harlow and Lane “Using Antibodies: ALaboratory Manual” Cold Spring Harbor Laboratory Press, 1999. The term“antibody” also comprises immunoglobulins (Ig's) of different classes(i.e. IgA, IgG, IgM, IgD and IgE) and subclasses (such as IgG1, IgG2etc.).

The definition of the term “antibody” also includes embodiments such aschimeric, single chain and humanized antibodies, as well as antibodyfragments, like, inter alia, Fab fragments. Antibody fragments orderivatives further comprise F(ab′)2, Fv, scFv fragments or singledomain antibodies, single variable domain antibodies or immunoglobulinsingle variable domain comprising merely one variable domain, whichmight be VH or VL, that specifically bind to an antigen or epitopeindependently of other V regions or domains; see, for example, Harlowand Lane (1988) and (1999), cited above. Such immunoglobulin singlevariable domain encompasses not only an isolated antibody singlevariable domain polypeptide, but also larger polypeptides that compriseone or more monomers of an antibody single variable domain polypeptidesequence.

As used herein, CD3 epsilon denotes a molecule expressed as part of theT cell receptor and has the meaning as typically ascribed to it in theprior art. In human, it encompasses in individual or independentlycombined form all known CD3 subunits, for example CD3 epsilon, CD3delta, CD3 gamma, CD3 zeta, CD3 alpha and CD3 beta. The human CD3epsilon is indicated in GenBank Accession No. NM_000733. The human CD19protein is indicated in GenBank Accession No. AAA69966.

Preferably, the bispecific antibody applied in the methods/dosageregimens of the present invention has the domain arrangementVL(CD19)-VH(CD19)-VH(CD3)-VL(CD3). It is, however, also envisaged thatthe methods of the invention can be carried out with CD19×CD3 bispecificsingle chain antibodies of other domain arrangements, such as

VH(CD19)-VL(CD19)-VH(CD3)-VL(CD3),

VL(CD19)-VH(CD19)-VL(CD3)-VH(CD3),

VH(CD19)-VL(CD19)-VL(CD3)-VH(CD3),

VL(CD3)-VH(CD3)-VH(CD19)-VL(CD19),

VH(CD3)-VL(CD3)-VH(CD19)-VL(CD19),

VL(CD3)-VH(CD3)-VL(CD19)-VH(CD19), or

VH(CD3)-VL(CD3)-VL(CD19)-VH(CD19).

A preferred CD19×CD3 bispecific antibody applied in the methods of thepresent invention comprises the

(a) anti-CD3 CDRs of the heavy chain shown as CD3 CDR-H1 in SEQ ID NO:11 (RYTMH), more preferably residues 26-35 of SEQ ID NO: 7 (GYTFTRYTMH),CD3 CDR-H2 in SEQ ID NO: 12 (YINPSRGYTNYNQKFKD) and CD3 CDR-H3 in SEQ IDNO: 13 (YYDDHYCLDY); and/or

(b) anti-CD3 CDRs of the light chain shown as CD3 CDR-L1 in SEQ ID NO:14 (RASSSVSYMN), CD3 CDR-L2 in SEQ ID NO: 15 (DTSKVAS) and CD3 CDR-L3 inSEQ ID NO: 16 (QQWSSNPLT); and/or

(c) anti-CD19 CDRs of the heavy chain shown as CD19 CDR-H1 in SEQ ID NO:17 (SYWMN), more preferably residues 26-35 of SEQ ID NO: 3 (GYAFSSYWMN),CD19 CDR-H2 in SEQ ID NO: 18 (QIWPGDGDTNYNGKFKG) and CD19 CDR-H3 in SEQID NO: 19 (RETTTVGRYYYAMDY); and/or

(d) anti-CD19 CDRs of the light chain shown as CD19 CDR-L1 in SEQ ID NO:20 (KASQSVDYDGDSYLN), CD19 CDR-L2 in SEQ ID NO: 21 (DASNLVS) and CD19CDR-L3 in SEQ ID NO: 22 (QQSTEDPWT).

It is more preferred that the CD19×CD3 bispecific antibody applied inthe methods of the present invention comprises the CD3 CDRs of the heavyand light chain. Even more preferably, the CD19×CD3 bispecific antibodyapplied in the methods of the present invention comprises the CD3 CDRsof the heavy and light chain as well as the CD19 CDRs of the heavy andlight chain.

The CDRs referred to herein are in accordance with the Kabat numberingsystem. The Kabat numbering scheme is a widely adopted standard fornumbering the residues in an antibody in a consistent manner (Kabat etal., Sequences of Proteins of Immunological Interest, 1991).

Alternatively, it is preferred that the CD19×CD3 bispecific antibodyapplied in the methods of the present invention comprises the

-   (a) CD19 variable heavy chain shown in SEQ ID NO: 3 (nucleotide    sequence is shown in SEQ ID NO: 4); and/or-   (b) CD19 variable light chain shown in SEQ ID NO: 5 (nucleotide    sequence is shown in SEQ ID NO: 6); and/or-   (c) CD3 variable heavy chain shown in SEQ ID NO: 7 (nucleotide    sequence is shown in SEQ ID NO: 8); and/or-   (d) CD3 variable light chain shown in SEQ ID NO: 9 (nucleotide    sequence is shown in SEQ ID NO: 10).

More preferably, the CD19×CD3 bispecific antibody applied in the methodsof the present invention comprises the CD19 variable heavy and lightchain and/or the CD3 variable heavy and light chain. Even morepreferably, the CD19×CD3 bispecific antibody applied in the methods ofthe present invention comprises the CD19 variable heavy and light chainas well as the CD3 variable heavy and light chain.

In another alternative, it is also preferred that said bispecific singlechain antibody comprises an amino acid sequence selected from the groupconsisting of

-   (a) an amino acid sequence as depicted in SEQ ID NO: 1;-   (b) an amino acid sequence encoded by a nucleic acid sequence as    shown in SEQ ID NO: 2;-   (c) an amino acid sequence encoded by a nucleic acid sequence having    at least 70%, 80%, 90%, 95% or 99% identity to a nucleic acid    sequence of (b), wherein said amino acid sequence is capable of    specifically binding to CD3 and CD19; and-   (d) an amino acid sequence encoded by a nucleic acid sequence which    is degenerate as a result of the genetic code to a nucleotide    sequence of (b), wherein said amino acid sequence is capable of    specifically binding to CD3 and CD19.

It is to be understood that the sequence identity is determined over theentire amino acid sequence. For sequence alignments, for example, theprograms Gap or BestFit can be used (Needleman and Wunsch J. Mol. Biol.48 (1970), 443-453; Smith and Waterman, Adv. Appl. Math 2 (1981),482-489), which is contained in the GCG software package (GeneticsComputer Group, 575 Science Drive, Madison, Wis., USA 53711 (1991). Itis a routine method for those skilled in the art to determine andidentify an amino acid sequence having e.g. 70%, 80%, 90%, 95%, 96%,97%, 98% or 99% sequence identity to the amino acid sequences of theCD19×CD3 bispecific antibody described herein (preferably MT103). Forexample, according to Crick's Wobble hypothesis, the 5′ base on theanti-codon is not as spatially confined as the other two bases, andcould thus have non-standard base pairing. Put in other words: the thirdposition in a codon triplet may vary so that two triplets which differin this third position may encode the same amino acid residue. Saidhypothesis is well known to the person skilled in the art (see e.g.,wikipedia.org/wiki/Wobble_Hypothesis; Crick, J Mol Biol 19 (1966):548-55). It is furthermore a routine procedure for those skilled in theart to determine cytotoxic activity of such an amino acid sequencehaving e.g. 70%, 80%, 90%, 95%, 96%, 97%, 98% or 99% sequence identityto the nucleotide or amino acid sequences of the CD19×CD3 bispecificsingle chain antibody described herein. Cytotoxic activity of theCD19×CD3 bispecific single chain antibody or an antibody constructhaving e.g. 70%, 80%, 90%, 95%, 96%, 97%, 98% or 99% sequence identityto the amino acid sequences of the CD19×CD3 bispecific single chainantibody can be determined by methods as illustrated e.g. in WO99/54440.

Particularly preferred, said CD19×CD3 bispecific single chain antibodyhas the amino acid sequence shown in SEQ ID NO: 1.

Also particularly preferred is the CD19×CD3 bispecific antibody MT103described in WO 99/54440 as well as those CD19×CD3 bispecific antibodiesdescribed in WO 2004/106381 and WO 2008/119565.

The present invention furthermore relates to a CD19×CD3 bispecificantibody for:

-   (i) administering a CD19×CD3 bispecific antibody to a human patient,    or-   (ii) treating malignant CD19 positive lymphocytes in a human    patient; and/or-   (iii) ameliorating or preventing an adverse effect mediated by the    administration of a CD19×CD3 bispecific antibody to a human patient;    wherein said antibody is to be administered in accordance with a    dosage regimen as defined in any one of the preceding disclosure.

Also, the present invention relates to a CD19×CD3 bispecific antibody

-   (i) administering a CD19×CD3 bispecific antibody to a human patient,    or-   (ii) treating malignant CD19 positive lymphocytes in a human    patient; and/or-   (iii) ameliorating or preventing an adverse effect mediated by the    administration of a CD19×CD3 bispecific antibody to a human patient;    wherein said antibody is to be administered in accordance with a    method as defined in any one of the preceding disclosure.

In a further aspect, the present invention concerns the use of aCD19×CD3 bispecific antibody for the preparation of a pharmaceuticalcomposition to be used in a method as defined in any one of the methodsdescribed herein.

The pharmaceutical composition of the present invention may optionallycomprise a pharmaceutical carrier. Examples of suitable pharmaceuticalcarriers are well known in the art and include phosphate buffered salinesolutions, sterile solutions etc. Intravenous vehicles include fluid andnutrient replenishers, electrolyte replenishers (such as those based onRinger's dextrose), and the like. Preservatives and other additives mayalso be present such as, for example, antimicrobials, anti-oxidants,chelating agents, and inert gases and the like. Furthermore, thepharmaceutical composition of the invention may comprise further agentssuch as chemotherapeutic agents as explained herein elsewhere.

In a further aspect, the present invention relates to a (pharmaceutical)kit or pharmaceutical package comprising the first dose and the seconddose as defined herein.

In another embodiment, the present invention relates to a(pharmaceutical) kit or pharmaceutical package comprising the first doseand the second dose as defined herein as well as the third dose asdefined in the context of the three-stage dosage regimen/method.

In another aspect, the (pharmaceutical) kit or pharmaceutical packagecomprises all three doses as defined herein in the context of thethree-stage dosage regimen/method, i.e., the first, the second and thethird dose.

Said first, second and third dose are thereby packaged together in onesealed pharmaceutical package or kit. It will be understood that the“first dose”, the “second dose” and the “third dose” encompasses in thisregard the respective number of single doses which will be used for agiven period of time (either the first or the second period of time).This means for example that the “first dose” or “second dose” which iscomprised in the pharmaceutical package or kit of the present inventioncomprises, for example, 7 daily doses which are separated. The number ofpackaged daily doses thereby reflects the intended period of time (Xdaily doses if said period of time is X days, Y daily doses if theperiod of time is Y days and so on). In these embodiments, the(pharmaceutical) kit or pharmaceutical package comprises the dailydosages in separate containers, in a single package.

Alternatively, it is also envisaged that the intended first dose and/orsecond dose and/or third dose is not separated into the respectivenumber of daily doses but is contained, either in toto or in part, inone single container (for example an infusion bag), which comprises therequired dose for either the first and/or the second period of timeeither in part (for example for 1 to 3 days) or in toto (i.e. for thefirst or second period of time). This means that one single containercomprises, for example, 7 daily doses for the “first dose” which is tobe used during the first period of time, and similarly for the secondand third doses.

It will be understood that the (pharmaceutical) kit or pharmaceuticalpackage of the present invention may also comprises more or less dailydoses as required for the respective period of time (either separated ornot). Alternatively, the (pharmaceutical) kit or pharmaceutical packageis prepared such that it contains the required number of daily doses(either separated or not) for the first and second period of time asdefined herein, i.e. the “first dose”, the “second dose” and the “thirddose” in one single package. Such a package is ideally sufficient forone complete treatment of a patient (including the first and the secondperiod of time). Parts of the kit and package of the invention can bepackaged individually in vials or bottles or in combination incontainers or multicontainer units. The manufacture of the kits followspreferably standard procedures which are known to the person skilled inthe art.

Furthermore, the invention relates to a pharmaceutical package or kit asdescribed hereinbefore and written instructions for the sequential usethereof in accordance with the methods of the present invention. Saidpharmaceutical package or kit may further comprise a label or imprintindicating that the contents can be used for treating malignant CD19positive lymphocytes present in lymphoma or leukemia in a human patient;or for ameliorating or preventing an adverse effect mediated by theadministration of a CD19×CD3 bispecific antibody to a patient.

It is also envisaged that the pharmaceutical package or kit of thepresent invention, further comprises means to administer the firstand/or the second dose and/or third dose to a patient and/or buffers,vials, teflon bags or infusion bags which are normally used for theinfusion of therapeutic agents. “Means” thereby includes one or morearticle(s) selected from the group consisting of a syringe, a hypodermicneedle, a cannula, a catheter, an infusion bag for intravenousadministration, intravenous vehicles, vials, buffers, stabilizers,written instructions which aid the skilled person in the preparation ofthe respective doses and infusions of the invention etc.

It is also envisaged that the pharmaceutical package or kit of thepresent invention further comprises a chemotherapeutic agent.

In a further aspect, the present invention provides for a pharmaceuticalpackage or kit, wherein said first and/or said second dose is arrangedsuch, that it is suitable for (prepared for) administration of a dosageregimen in accordance with a method of any one of the precedingdisclosures.

THE FIGURES SHOW

FIG. 1: B cell counts from patient samples are stratified as predictorof risk of adverse effects in patients treated with a CD19×CD3bispecific antibody.

FIG. 2: B cell counts of patient samples as a predictor in comparison toB:T cell ratios. A) and B) are results from two clinical studiestreating patients with a CD19×CD3 bispecific antibody (A: trial 103-104,NHL; B: trial 103-202, ALL).

EXAMPLES

The following examples illustrate the invention. These examples shouldnot be construed as to limit the scope of this invention. The examplesare included for purposes of illustration and the present invention islimited only by the claims.

Example 1

Identification of a predictive factor for reversible neurologicaladverse events in a subset of non-Hodgkin lymphoma patients treated withCD19-specific BiTE antibody blinatumomab

Blinatumomab is a CD19/CD3-bispecific antibody construct of thebispecific T cell engager (BiTE®) class showing as single agent a highrate and duration of responses in patients with relapsed non-Hodgkinlymphoma (NHL) and B-precursor acute lymphocytic leukemia (ALL).Blinatumomab has a favorable safety profile with exception of a subsetof patients developing neurological adverse events (AEs) during thefirst days of treatment, such as confusion, speech impairment orcerebellar symptoms. Thus far, all relevant neurological AEs (11 out of48 patients) were transient, fully reversible and resolved withoutsequelae within 3 to 72 hours after stop of infusion. In no case,pathological findings were seen upon cranial magnetic resonance imaging.Despite treatment discontinuation, 4 patients with neurological AEs haveachieved an objective lymphoma remission. Analysis of cerebrospinalfluid (CSF) taken within hours after stop of infusion showed detectablelevels of blinatumomab in the majority of affected patients, while inone patient without neurological symptoms no blinatumomab was detectablein CSF during infusion. Moreover, increased levels of albumin and Tlymphocytes in CSF support a disturbance of the blood brain barrier(BBB) as a possible underlying event. Analyses of patient serum samplesfor angiopoetin-2 and S100β are ongoing to investigate whether levels ofthe endothelial stress and BBB integrity marker, respectively, correlatewith neurological AEs. In a retrospective analysis of 39 NHL patients, abaseline B cell to T cell (B:T) ratio in peripheral blood at or below1:10 was identified as the only predictive factor for the subsequentoccurrence of neurological AEs. The predictive value was thenprospectively confirmed in 8 additional patients. In conclusion, asimple measure to prospectively identify patients at risk of developingneurological AEs after onset of blinatumomab treatment has beenidentified. Mitigating measures are currently being tested in thesehigh-risk patients in order to avoid discontinuation of treatment.

Example 2

Synopsis of observations (1) in patients treated with a CD19×CD3bispecific antibody

Common Features of Early CNS Events

-   -   First symptoms appear 12-48 h after start of MT103 infusion:        Agitation, speech impairment, sometimes tremor, ataxia    -   More severe symptoms leading to Infusion stop appear after 24-72        h: Confusion, disorientation, ataxia, aphasia, seizure    -   After stop of MT103 infusion, complete resolution of CNS        symptoms seen within 1-3 days; generally no sequelae    -   Most CNS events fall into early activation and redistribution        phase of polyclonal T cells

Features of CNS Events with Slow Onset

-   -   Biased to cerebellar symptoms    -   Occur at various time points during treatment, frequently at        beginning of treatment or at step increase    -   Tremor, mild speech impairment, mild writing impairment; can        last for several days

Other CNS Events

-   -   Additional symptoms observed without proven relationship to        other CNS events: Headache, Fever, Nausea

MT103 Dose Response Relationship of CNS Events

-   -   Dose response relationship of CNS events is evident; cut off is        between dose level of 5 and 15 μg/m²/d

Example 3

Synopsis of observations (2) in patients treated with a CD19×CD3bispecific antibody

CNS Events Appear to be Predictable

-   -   Correlation of CNS events with low B:T cell ratio (or low B cell        count)    -   B:T ratio of <1:10 identified as apparent cut off for        development of CNS events    -   No other biochemical or clinical parameters appear to correlate        with CNS events

Cranial MRI of Patients with CNS Events Mostly without PathologicalFindings CSF Analyses Suggest Opening of BBB and Neuroinflammatory Event

-   -   Detectable levels of MT103 and increased levels of protein and        serum albumin found in majority of affected patients suggest        temporary breakdown of blood brain barrier (BBB)    -   No MT103 found in CSF of one patient free of CNS events    -   CSF analysis also shows in some affected patients increased        counts of monocytes and T lymphocytes indicative of        neuroinflammatory process    -   Are CNS events reflecting gradual opening of BBB        (agitation>confusion>aphasia, ataxia>seizure)?

Incidence of CNS Events May Correlate with Disease and/or Tumor Load

-   -   At 15 μg/m²/d, 3/8 NHL patients (37%) and only one of 1/11 ALL        ‘high risk’ patients (9%) developed CNS events    -   B-ALL patients routinely receive intrathecal chemotherapy (and        i.v. high-dose methotrexate) likely reducing tumor cell load in        CNS (“occult meningeosis neoplastica”)

Example 4

Summary of CNS events in patients treated with a CD19×CD3 bispecificantibody

Summary of Clinically Relevant CNS Events in NHL Patients First or DoseTreatment Complete Neurological B:T cell Additional in Stop afterResolution, Best Patient # Assessment Disease ratio Gender, AgeTreatment μg/m²/Day Start Time Response 105-005 Confusion, FL  1:23.9Female, 65 First 15 15 h Yes, 24 h SD communication disorder 102-004Organic Brain MCL 1:757 Male, 75 First 15 50 h Yes, 34 h n.d. Syndrome102-006 Generalized MZL  1:1740 Male, 59 First 30 48 h Yes, 48 h n.d.seizure (acidosis) 109-011 Cerebellar MCL 1:9.2 Male, 73 Restart 60 48 hYes, 24 h PR Symptoms (first) 109-012 Encephalopathy MCL  1:19520 Male,55 Additional 60 24 h Yes, 24 h CR (first) 102-007 Seizure, aphasia FL1:197 Male, 61 First 90 48 h Yes, 48 h ?PR? 109-023 Encephalopathy MCL1:368 Male, 60 First 60 17 h Yes, 56 h n.d. 109-025 Encephalopathy MCL1:873 Male, 58 First 15 41 h Yes, 48 h n.d. 108-004 Speech FL 0:431Male, 66 First 60 624 h  Yes, 3 h PR Impairment Palsy Face and Arm109-261 Desorientation, MCL 1:20  Male, 42 Additional 60 30 h Yes, 72 hPR Speech (first Impairment cycle)

Example 5

Dose dependency of CNS events of patients treated with a CD19×CD3bispecific antibody in Clinical Trials

Dose Dependency of CNS Events in Ongoing NHL Trial ‘High risk’ patientsdefined by having low B:T cell ratio (<1:10) Initial dose considered forclassification in dose groups Dose All ‘High Risk’ ‘Low Risk’ ≤5 0/14(0%) 0/4+ (0%) 0/10 (0%) 15 3/16 (19%) 3/8+ (38%) 0/8 (0%) 30 1/6 (17%)1/1 (100%) 0/5 (0%) 60 5/13 (38%) 4/5 (80%) 1*/8 (13%) 90 2/3 (66%) 1/1(100%) 1/2 (50%) All 11/52^(§) (21%) 9/19 (47%) 2/33 (6%) ^(§)>48patients is due to additional treatments and re-starts of individualpatients (resulting in conversion to ‘high risk’) *Reached borderlineB:T ratio after first treatment cycle +Incl. patients with step-wisedose increase

Example 6

A patient having an increased risk of potential adverse effects whoreceived 15 μg/m²/d for 7 days and 60 μg/m²/d for 21 days showed noadverse effects (neurological reactions)

Patient 108-003

-   -   Female, 66 y    -   FL grade 2, IVB (FD: 09/2006)    -   Relevant medical history: anemia, thrombocytopenia,        (pre-treatment 2× Zevalin and bone marrow infiltration by FL)        elevation of gGT and AP, abuse of benzodiazepines, status        after 2. aureaus sepsis with spondylodiscitis and abscesses    -   Prior lymphoma treatment:        -   6×R-CHOP 14, 8×R 09/2006-02/2007        -   R mono 05/07        -   1. Zevalin 11/07        -   2. Zevalin 01/08    -   According to initial B:T cell ratio (1:10.5) high-risk (cohort        15/60)    -   Jan. 5, 2009 Treatment start (15 μg/m²/24 h)    -   Fever, headache for 2 days—easily handled by oral paracetamol        and novalgin    -   January 12th dose increase to 60 μg/m²/24 h    -   Again fever, headache—easily handled by oral paracetamol and        novalgin    -   No neurological events    -   Well tolerated dose “step”    -   Suspected improvement of bone marrow function

Example 7

A patient having an increased risk of potential adverse effects whoreceived 5 μg/m²/d for 7 days and 60 μg/m²/d for 21 days showed mildadverse effects (neurological reactions)

-   -   MCL, male 42 y    -   B:T 1:12    -   Treatment start Jan. 19, 2009 with 5 micg/m2/d    -   Day 1: fever and chills, headache, no further problems    -   Step: January 26^(th): after 6 h fever, strong headache    -   27 Jan. 2009: tiredness, nausea, vomiting, endoscopy without        pathological findings), absolute arrhythmia with frequency up to        170/min→resolution within one day after substitution of        potassium and digitoxin.    -   Cranial CT scan and CSF performed, CT: no pathological findings    -   CSF: slightly elevated protein 55 mg/dL, cells: 23 Zellen/micL,        mainly monocytic cells and some activated lymphocytes    -   27 Jan. 2009 afternoon: mild tremor, apraxia, “slow mental        state”, evening: mild speech impairment (cerebellar?), slow        improvement over the next two days 29 Jan. 2009 due to ongoing        mild symptoms decision to give dexamethasone    -   Slow improvement of symptoms, complete resolution 31 Mar. 2009    -   During the further course of treatment: recurrent difficulties        to play the guitar.    -   After 4 weeks treatment: −37%    -   After 8 weeks of treatment: PR/CRu

Example 8

A patient having an increased risk of potential adverse effects whoreceived a treatment regimen according to the present invention.

Patient 108-005

-   -   Male, 71 y, FL IIIB    -   B:T cell ratio: 57:1363 (low, 1:23.9)    -   First diagnosis: 1997    -   Multiple prior treatments: 12× Rituximab (mono), 6×        Rituximab-Bendamustin, 6×R-CHOP, autologous SCT    -   Date of Blinatumomab start: 17 Oct. 2009    -   Treatment duration: 8 weeks    -   Well tolerated (no SAE)    -   No neurological adverse event    -   8 Week CT Scan: −65%=partial remission of the lymphoma

Example 9

A further patient having an increased risk of potential adverse effectswho received a treatment regimen according to the present invention.

Patient 109-031

-   -   Male, 60 y, Follicular Lymphoma IVAE    -   B:T cell ratio: 0:429 (low)    -   First diagnosis: 05/09    -   Prior treatments: Pre-phase w. Vincristin/Decortin, 6×R-CHOP    -   Blinatumomab treatment Start: 30 Nov. 2009    -   Treatment duration: 8 weeks    -   Well tolerated (flush symptoms at steps—responsive to steroids)    -   No neurological adverse event    -   Lymphoma—56% after 8 weeks (partial remission of the lymphoma)

The invention claimed is:
 1. A method for treating a human patientsuffering from malignant CD19-positive lymphoma or leukemia, wherein thepatient is at risk for potential adverse effects resulting fromtreatment with a CD19×CD3 bispecific antibody comprising an amino acidsequence encoded by the nucleic acid sequence set forth in SEQ ID NO: 2or the amino acid sequence set forth in SEQ ID NO: 1, the methodcomprising: (a) determining in the peripheral blood of the patient aratio of B cells to T cells and total B cell count; and (b) identifyingthe patient at risk as comprising a ratio of B cells to T cells of about1:8 or lower; (i) wherein a total B cell count of less than 40 B cellsper microliter of peripheral blood is indicative of a higher risk forpotential adverse effects for said patient; and (ii) wherein a total Bcell count of 40 B cells or greater per microliter of peripheral bloodis indicative of a lower risk of potential adverse effects for saidpatient, (c) administering the CD19×CD3 bispecific antibody (i) to thepatient who is at higher risk in an incremental dosage schedule toreduce the possibility of potential adverse effects in the patient,wherein a first dose is administered for a first period of time and asecond dose is consecutively administered for a second period of time,wherein said second dose is greater than said first dose, and whereinsaid second period of time exceeds said first period of time; or (ii) tothe patient who is at lower risk in a constant dose for at least fourweeks.
 2. The method of claim 1, wherein the specific dosage schedulefor said higher risk patient comprises: (a) administering a first doseof the CD19×CD3 bispecific antibody for a first period of time of 7days; and consecutively (b) administering a second dose of said antibodyfor a second period of time that exceeds the first period of time; andoptionally consecutively, (c) administering a third dose of saidantibody for the first or second period of time or for a third period oftime, wherein said second dose exceeds said first dose and said thirddose exceeds said second dose.
 3. The method of claim 1, wherein saidadverse effect is characterized by a neurological reaction.
 4. Themethod of claim 2, wherein: said second period of time exceeds saidfirst period of time; said third period of time exceeds said firstperiod of time and/or said second period of time; said first period oftime exceeds 3 days; said first period of time is between 3 and 10 days,or is 7 days; said second period of time exceeds 18 days; said secondperiod of time is between 18 and 81 days, is 21 days, or is 49 days;said first period of time is between 3 days and 10 days, and said secondperiod of time is between 18 and 81 days; or said first period of timeis 7 days and said second period of time is 21 days or is 49 days. 5.The method of claim 2, wherein said first dose is between 1 and 15μg/m²/d, or 5 or 15 μg/m²/d; or said second dose is between 15 and 60μg/m²/d, or is 60 μg/m²/d.
 6. The method of claim 2, further comprisingadministering a third dose of said antibody for a third period of timeafter a first and second dose for a first and second period of time. 7.The method of claim 6, wherein said third period of time exceeds saidfirst and second period of time, and wherein said second dose exceedssaid first dose; said first period of time exceeds 3 days; said firstperiod of time is between 3 and 10 days, or is 7 days; said secondperiod of time exceeds 3 days; said second period of time is between 3and 10 days, or is 7 days; said third period of time exceeds 8 days;said third period of time is between 8 and 78 days, or is 14 days, or is42 days; said first period of time is between 3 and 10 days, and saidsecond period of time is between 3 and 10 days, and said third period oftime is between 8 and 78 days; or said first period of time is 7 days,said second period of time is 7 days, and said third period of time is14 days, or is 42 days.
 8. The method of claim 6, wherein said thirddose exceeds said first and second dose; said first dose is 5 μg/m²/d oris between 1 and 15 μg/m²/d, or wherein said first dose is between 2 and29 μg/d, or is 9 or 10 μg/d; said second dose is 15 μg/m²/d or isbetween 1 and 15 μg/m²/d, or wherein said second dose is between 2 and29 μg/d, or is 28 or 29 μg/d; or said third dose is 60 μg/m²/d or isbetween 15 and 60 μg/m²/d, or wherein said third dose is between 28 and116 μg/d, or is 112 μg/d.
 9. The method of claim 1, wherein the lymphomacomprises indolent or aggressive B cell non-Hodgkin lymphoma (B NHL),mantle cell lymphoma (MCL) or chronic lymphatic leukemia (CLL).
 10. Themethod of claim 1, wherein the leukemia comprises B-lineage acutelymphoblastic leukemia (ALL).
 11. The method of claim 1, furthercomprising periodically determining the ratio of B cells to T cellsand/or total B cell counts during administration of said antibody. 12.The method of claim 11, wherein the administering of the bispecificantibody may be adjusted based upon results of periodically determiningthe ratio of B cells to T cells and/or total B cell count to reduce therisk for potential adverse effects during the treatment.
 13. The methodof claim 1, wherein said second period of time exceeds 18 days.
 14. Themethod of claim 13, wherein said second period of time is between 18days and 81 days, or between 21 and 49 days.
 15. The method of claim 13,wherein said first period of time is 7 days and said second period oftime is 21 or 49 days.
 16. The method of claim 1, wherein said firstdose is between 1 and 15 μg/m²/d, or wherein said first dose is between2 and 29 μg/d.
 17. The method of claim 1, wherein said second dose isbetween 15 and 60 μg/m²/d, or wherein said second dose is between 28 and116 μg/d.
 18. The method of claim 1, wherein the method furthercomprises administering a third dose of said antibody for a third periodof time after a first and second dose for a first and second period oftime.
 19. The method of claim 18, wherein said third period of timeexceeds said first and second period of time, and wherein said seconddose exceeds said first dose.
 20. The method of claim 18, wherein saidthird dose exceeds said first and second dose.